Impact of a temporal salinity decrease on growth and nitrogen metabolism of the marine diatom Skeletonema costatum in continuous cultures

In 1987 effects of salinity fluctuations on growth of the centric diatom Skeletonema costatum (Greville) Cleve, isolated from the brackish Krammer estuary (SW Netherlands) in 1981, were investigated. Continuous cultures (12 h light: dark cycle) of S. costatum were adapted to constant salinity in natural (16.1) and synthetic (13.5) media. For several days the ammonium-limited cultures were exposed to a salinity fluctuation (minimum 4.8). Decreasing salinity caused an inhibition of photosynthesis, dark respiration and cell growth. Cellular pools of glucose decreased. While the carbohydrate content remained constant, the protein content increased slightly. Net carbon fixation was more inhibited than nitrogen assimilation. Ammonium accumulated during a salinity decrease; a total decline of the overcapacity of ammonium uptake was noticed and nitrogen limitation was relieved. Amino acid pools decreased, probably as a result of excretion (osmoregulation). The enzymes invoilved in ammonium assimilation showed an increased activity. Cellular activities were resumed during a salinity increase. Chlorophyll a increased; photosynthesis, ammonium uptake and growth were stimulated. The ammonium uptake capacity recovered completely; glutamic acid accumulation and increased glutamate-dehydrogenase (GDH) activity indicated supplementary ammonium assimilation via GDH. The activities of glutamine synthetase/glutamate synthase (GS/GOGAT) and GDH stabilized, and the cells returned to steady state under ammonium limitation.Communication no. 426 Delta Institute for Hydrobiological Research, Yerseke, The Netherlands     

Aspects of nitrogen metabolism of the common mussel Mytilus edulis: Adaptation to abrupt and fluctuating changes in salinity

Mytilus edulis L. were exposed to abrupt (3015 and 1530) and fluctuating (sinusoidal 12 h cycles of 301530) changes in salinity, and the changes in the total osmoconcentration of the haemolymph were recorded. The response of nitrogen metabolism to the altered extracellular osmotic concentrations was investigated in terms of the concentrations of the total NPS (ninhydrin-positive substances) pool and the individual amino acids of the tissues, the concentration of the amino acids of the haemolymph, and the rates of excretion of ammonia and amino acids by whole individuals. The haemolymph became isosmotic with the seawater with abrupt changes in salinity, but with fluctuating salinity was slightly hyperosmotic as the salinity decreased and then slightly hypo-osmotic as the salinity increased. This resulted in a reduction in the extent of the extracellular osmotic change compared to the change in fluctuating salinity to which it was exposed. Total NPS of the tissues decreased with an abrupt decrease in salinity and increased with an abrupt increase in salinity, but a seasonal dependence of the response was indicated. The short-term response of tissue NPS to fluctuating salinity was equivocal, but with long-term exposure the concentration declined. Ammonia and amino acid excretion increased with both an abrupt decrease in salinity and fluctuating salinity and decreased with an abrupt increase in salinity. Haemolymph amino acids increased with an abrupt decrease in salinity. The increased rates of nitrogen excretion accounted for the reductions in the NPS concentrations of the tissues except in the early stages of fluctuating salinity. Taurine, aspartate, threonine, serine, glycine and arginine declined with an abrupt decrease in salinity while alanine and glutamate increased slightly. With an abrupt increase in salinity, alanine and ammonia accumulated in the tissues and then declined while the other amino acids increased slowly over a longer time-course. Similar individual amino acid responses were seen with long-term exposure to fluctuating salinity, except for taurine which did not decrease in concentration. On the basis of the changes in tissue amino acids and ammonia, it is suggested that the alanine dehydrogenase reaction is the primary nitrogen-fixing reaction in marine bivalves such as M. edulis.     

HPLC analysis of nonprotein thiols in planktonic diatoms: pool size,redox state and response to copper and cadmium exposure

A sensitive method was developed to analyze low molecular weight thiols involved in metal homeostasis and detoxification in phytoplankton. The aims of this study were to (1) separate and measure all relevant thiols in a single HPLC run, (2) measure redox states of the thiols and (3) identify specific responses of thiols (pools, redox) to heavy metals by testing diatoms with different metal tolerances (Ditylum brightwellii, Phaeodactylum tricornutum, Skeletonema costatum andThalassiosira pseudonana). Copper or cadmium were dosed at a maximum tolerable, species-dependent level, to exponential phase cells growing in artificial medium (14 salinity). Loss of cell viability was monitored by the decrease of fluorescein fluorescence after a 24-h metal exposure. Thiols in extracts of exposed cells and controls were labeled with monobromobimane. Picomoles of cysteine, glutathione, -glutamylcysteine and phytochelatins (PCs) were detected and separated by reversed-phase high performance liquid chromatography. Cysteine increased in all species after metal exposure. The species with the largest glutathione pools in the control (P. tricornutum) synthesized the largest PC pools upon metal exposure, however, at a 40% loss of glutathione. A considerable increase of glutathione was observed inD. brightwellii upon metal exposure. However, it produced little PC (and only with Cu). In controls ( 3 pM Cu²⁺), PC₂ was detectable inS. costatum, P. tricornutum andT. pseudonana. Oxidized thiol fractions were recovered by the reductants DTT and TCEP, both performing identically. Compared to the other thiols, cysteine had low redox ratios. InD. brightwellii glutathione and PC redox ratios were lower than inP. tricornutum, S. costatum andT. pseudonana. It was expected that Cu-induced oxidative stress would decrease the thiol redox ratios, however, this was not observed.This is Communication No. 2172 of NIOO.     

Effects of salinity and temperature on Corophium volutator and C. arenarium (Crustacea: Amphipoda), with particular reference to distribution

The responses of the post-embryonic stages of Corophium volutator (Pallas) and C. arenarium Crawford to the combined effects of salinity and temperature show that gravid females have a wider tolerance than nongravid adult females which in turn are more tolerant than adult males. C. volutator is more tolerant of low salinity (2 to 10) than C. arenarium, but the latter is more tolerant of salinities above 45. The embryos of C. volutator develop normally and hatch at lower salinities and temperatures than those of C. arenarium, in which successful development was recorded at higher temperatures. Females undergoing a pre-copulatory moult failed to lay eggs below salinities of 3 (C. volutator) and 10 (C. arenarium), but in both species the lowest salinity at which all females moulted and laid eggs was 20. The results are discussed in relation to the distribution of both species.     

Influence of salinity on embryonic development and the distribution ofSepia officinalis in the Delta Area (South Western part of The Netherlands)

The effect of salinity on embryonic development ofSepia officinalis (cuttlefish) in the Delta Area (South Western part of The Netherlands) was investigated in 1988/1989, and compared with data concerning the distribution ofS. officinalis in the three main parts of this area: Oosterschelde, Westerschelde and Grevelingen. Embryos hatched in water collected at Yerseke (Oosterschelde), Vlissingen (Western part of the Westerschelde) and Bommenede (Grevelingen), i.e., at salinity values above 28.1, but not in water sampled at Hoedekenskerke and Hansweert (Middle and Eastern part of the Westerschelde; salinities below 22.0). Under laboratory conditions, using diluted Oosterschelde water, the highest hatching percentages ofS. officinalis were found at salinities above 29.8. Some embryos hatched at a salinity value of 26.5 but no hatching occurred at salinities below 23.9. In embryos exposed to salinity changes during late embryonic development, the developmental rate decreased at salinity values of 28.7 or less. Below 22.4 embryos with morphological malformations were found. It can be concluded that salinity is an important factor limiting the distribution ofS. officinalis in most parts of the Delta Area, with the exception of the Western part of the Westerschelde and the Grevelingen.Contribution no. 489 of the Library of the Delta Institute for Hydrobiological Research     

Kinetics of osmoregulation in the crab Carcinus maenas

When shore crabs Carcinus maenas are transferred from 11 to 38 S at 11°C, new constant levels of hemolymph freezing points and of concentrations of Na, K, Ca, and Mg in the hemolymph are accomplished within 24h. From a decrease in serum protein and in serum free amino acids and an increase in the relative amounts of individual essential free amino acids in the body fluids of whole crabs, a participation of proteolytic activities and a transport of amino acids from hemolymph to cell is deduced; the stationary concentration of total intracellular free amino acids increases up to a nearly two-fold value, compared with the concentration in crabs remaining in diluted sea water. Also, the low molecular neutral sugars increase in whole crabs after high salinity stress, reaching values of more than two-fold initial concentrations within a period of 10 days. This increase is fully accounted for by a 6.7-fold increase in trehalose levels. The processes of increasing the concentrations of low molecular organic material seem to be slower than those of establishing new osmotic pressures in the hemolymph. The oxygen consumption decreases by 30 to 45% to new constant values within 8 to 12 h after the salinity change from 11 to 38, reflecting similar kinetics as the establishment of new osmotic pressures in the hemolymph.     

Effects of salinity and calcium concentration on arm regeneration byOphiothrix angulata (Echinodermata: Ophiuroidea)

At 33 salinity a tissue stump formed 2 to 3 d after autotomy and developing ossicles were present by the fourth day inOphiothrix angulata (Say). Regeneration proceeded rapidly from the sixth day until the thirteenth day, when the rate decreased greatly. The length of the regenerated arm and the number of ossicles formed did not vary over a salinity range of 28 to 38 S, but were significantly less at 23 S. The number of ossicles regenerated increased linearly (y=1.9 x-7.7;r=0.9089) with the calcium concentrations ranging from 3.8 to 9.5 mM. No ossicle formation occurred at 3.8 mM calcium concentration. Rate of net uptake of calcium-45 into the ossicles of intact individuals in salinities of 28 and 33 was significantly greater than that in 23 and 38 S. However, net uptake rate of calcium into the soft tissues of the arms was significantly higher at 18S than at the lower two salinities.     

Responses of Leptasterias hexactis (Echinodermata: Asteroidea) to low salinity

The six-rayed starfish Leptasterias hexactis (Stimpson, 1862) is seasonally exposed to low salinities in southeastern Alaska. Individuals that were gradually exposed to reduced salinities in the laboratory had a 28-d TLm of 12.9 S. The activity of L. hexactis, as measured by its activity coefficient, varied directly with salinity. Individual feeding rates of the starfish on similarly exposed Mytilus edulis, measured daily for 21 d at salinities of 30, 20 and 15 S, also varied directly with salinity. The dry weight of mussel tissue consumed was 8.84, 8.49 and 0.58 mg ·starfish^-1·d^-1 at 20, 20 and 15 S. Expressed as percent of dry starfish weight, the daily feeding rate was 1.35, 0.76 and 0.10% at 30, 20 and 15 S. Absorption efficiency decreased from 64% at 30 S to 49% at 20 S, further reducing the energy available for metabolism. Growth, measured in terms of changes in total dry weight or dry weight of soft tissues, also varied directly with salinity. Although exposure to hyposmotic conditions did induce stress responses, as indicated by reductions in activity, feeding, absorption efficiency and growth rates, L. hexactis maintained positive growth for at least a 3-wk period in the laboratory at 20 S and 13°CC. The population of L. hexactis investigated must be considered euryhaline and brief periods of exposure to hyposmotic conditions should not limit its distribution.     

Effects of temperature,nutritional factors and salinity on the uptake of L-methionine by the Sydney rock oyster Saccostrea commercialis

The effects of physiological and nutritional factors and of temperature on the uptake of L-methionine by Saccostrea commercialis were investigated on cultured rock oysters from Port Stephens, New South Wales, Australia, in 1983. Optimum conditions for L-methionine accumulation were 30°C, 18 h exposure and concentrations of other amino acids less than that of L-methionine. The uptake of L-glycine was inhibited by L-methionine in a reciprocal manner. There was no effect of salinity on the accumulation of L-methionine by acclimated oysters. During the latter investigations the range of osmoconformity was found to be 15 to 45 S. Oysters take 2 d to conform to new media osmolarities after salinity changes of 15. Amino acid supplements in oyster diets should be kept within the same order of magnitude to reduce inhibition of uptake.     

Effects of salinity fluctuations on the respiration rate of the southern oyster drill Thais haemastoma and the blue crab Callinectes sapidus

Respiration rates of Thais haemastoma and Callinectes sapidus were determined as a function of salinity with a flow-through respirometer at 20°C. Respiration rates were measured at 10, 20 and 30 S for acclimated animals. The effects of 10-5-10, 20-10-20, 30-10-30 and 10-30-10 S semidiurnal cycles (12 h) of fluctuating salinity on the rate of respiration of the oyster drill were studied. During each cycle, salinity was changed from the acclimation salinity over a 4 h interval, held at that salinity for 2 h, returned to the acclimation salinity over 4 h and held at that salinity for 2 h. The effects of diurnal (24.8 h) salinity cycles on respiration in the oyster drill and blue crab were also studied. Salinity was changed from the acclimation salinity over a 10.4 h interval, held at that salinity for 2 h, then returned to the acclimation salinity over 10.4 h and held at that salinity for 2 h. The respiration rate of 30 S acclimated oyster drills (679 l O₂ g dry weight^–1 h^–1) was significantly higher than for individuals acclimated to 10 S (534 l O₂ g dry weight^–1 h^–1). Blue crab respiration was 170 l O₂ g dry weight^–1 h^–1 at 30 S, and was significantly higher at 10 and 20 S than at 30 S. With the exception of the 20-10-20 S semidiurnal cycle, the respiration rate of oyster drills declined as salinity fluctuated in either direction from the acclimation salinity and increased as ambient salinity returned to the acclimation salinity. Semidiurnal cycles (12 h) of fluctuating salinity produced greater changes in the respiration rate of snails than analogous diurnal cycles (24.8 h). A 10-30-10 S pattern of fluctuation caused a greater percentage reduction in the steady state respiration rate of oyster drills than the 30-10-30 S pattern. The respiration rate of blue crabs varied inversely with fluctuating salinity. Relatively minor changes occurred in blue crab respiration rate with fluctuating salinity. Blue crab respiration rate characteristically dropped during the initial phase of declining salinity at a rate directly proportional to the rate of salinity decrease, perhaps representing a metabolic adjustment period by the blue crabs. The respiratory response of T. haemastoma to salinity is consistent with its incomplete volume regulation, while the response of C. sapidus is compatible with its ability to regulate extracellular fluid osmotic and ionic composition.     

Salinity dependence of sexual and asexual reproduction in the rotifer Brachionus plicatilis

A salinity dependent mictic response was observed in a clone of Brachionus plicatilis cultured in the 2 to 4 salinity range. This response was related to asexual exponential reproduction rates (G) and could be divided into three categories: (a) no mixis occurred at a salinity of 35 S and above, where G values were lower than 0.30 d^-1, (b) low mictic levels in rotifers cultured at 2 and 30 S, where G values ranged between 0.40 to 0.50 d^-1, and (c) high mictic levels in rotifers cultured at salinities ranging between 4 and 20 S, where G values ranged between 0.50 to 0.85 d^-1. Fluctuations in mictic levels varied with time during the course of the experiments. Results suggest that salinity conditions leading to optimal parthenogenic reproduction also support mixis.     

Stepwise acclimation—a method for estimating the potential euryhalinity of the gastropod Hydrobia ulvae

The White Sea gastropod Hydrobia ulvae (Pennant) was exposed to step-wise lowering or increase of the habitat salinity. The time allowed for acclimatization to the successive salinity levels was sufficient to complete non-genetic adaptation. In this way, the lower and upper salinity limits were extended. The tolerance limits obtained are assumed to be indicative of the capacity for non-genetic adaptation and to serve as a genotypical characteristic. The tolerance of specimens colleced from in situ conditions (mid littoral, 20 S) ranged between 14 and 34 S. After non-genetic adaptation, the lower tolerance value shifted to 6 S (adaptation limit), and the upper value to 76 S (final limit not reached). There is no reason for considering White Sea H. ulvae to represent a special physiological race of specimens from those on the coast of Great Britain.     

The significance of temperature,salinity and zinc as lethal factors for the mussel Mytilus edulis in a polluted estuary

Mussels, Mytilus edulis L., were subjected to high temperatures, low salinities and dissolved zinc in order to investigate possible environmental hazards of a discharge of heated effluent near Newport on the Yarra River estuary, Victoria, Australia. Exposure to zinc at 0.8 mg l^-1 for 14 d in otherwise favourable conditions significantly increased mortality resulting from subsequent exposure to temperatures between 29° to 31°C for 24 h without added zinc. Mussels collected from water of temporarily lowered salinity (8–16 S) showed significantly lower thermal resistance than controls collected from marine salinities (35 S). Mussels taken from a marine environment and exposed to 10 S died at a rate which increased with temperature. Mussels acclimated for 14 d to combinations of 10°, 16° and 22°C and 22 and 35 S, and subsequently exposed to increased zinc concentrations accumulated zinc to levels which were independent of temperature and salinity. The zinc was lethal more quickly at 22°C and 35 S than at the lower temperatures and salinities. The modes of toxic action of the salinity, zinc and temperature factors are discussed and it is argued that zinc which has been found accumulated in mussels near Newport could be reducing their resistance to raised temperatures and perhaps other stresses, probably as a result of effects on lysosomal functioning. The evidence suggests that the heated effluent will accelerate any toxic effects of zinc or low salinities which occur near Newport and so poses a hazard in winter as well as in summer.     

Salinity tolerance,salinity preference and temperature tolerance in the high-shore harpacticoid copepod Tigriopus brevicornis

Tigriopus brevicornis (O. F. Müller) were collected in 1992 from rock pools close to U.M.B.S. Millport, Isle of Cumbrae, U.K. and acclimated to various combinations of salinity and temperature for at least 1 wk prior to laboratory experiments. Higher salinities of acclimation enhanced tolerance to high salinity stress, while tolerance of low salinities was hardly affected by acclimation salinity. Acclimation to low temperature (10°C) extended the survivable salinity range for T. brevicornis. High-salinity acclimation enhanced the survivable temperature range. Copepods acclimated to 60 survived significantly lower and higher temperatures than did 34-acclimated individuals. At high temperature, 75-acclimated female copepods had the highest median lethal temperature, 38.9°C. Females were significantly more resistant to high temperatures than males. The copepods were seen to have a very low median lethal temperature when frozen into solid ice for 2 h; 50% mortality occurred at-16.9°C in 10°C, 34-acclimated T. brevicornis. Salinity preference experiments demonstrated an ability to discriminate between salinities differing by as little as 3. Copepods acclimated to 34 chose salinities near their acclimation salinity; individuals acclimated to 5 favoured slightly higher salinities, while copepods acclimated to 60 chose rather lower salinities.     

Effects of salinity on respiration and nitrogen excretion in two species of echinoderms

The 30-d survival limit of Eupentacta quinquesemita and Strongylocentrotus droebachiensis is 12–13 S. The activity coefficient (1 000/righting time in seconds) of stepwise acclimated sea urchins declined from 16.3 at 30 S to 3.5 at 15 S. Oxygen consumption rates (QO₂) of both species held at 30 S and 13°C were highest in June and lowest in December. During the summer, when environmental salinity is most variable in southeastern Alaska, the QO₂ of both species held at 30, 20 and 15 S varied directly with salinity. Perivisceral fluid PO₂ varied directly with acclimation salinity in sea urchins, but not in sea cucumbers. Perivisceral fluid oxygen content of acclimated sea urchins was significantly lower at 15 and 20 S than at 30 S due to reduced PO₂ and extracellular fluid volume at the lower salinities. The QO₂ of both species varied directly with ambient salinity during a 30-10-30. semidiurnal pattern of fluctuating salinity. No change occurred in the average QO₂ of either species over a 15-30-15. semidiurnal pattern of fluctuating salinity. Sea urchin perivisceral fluid PO₂ declined as ambient salinity fluctuated away from the acclimation salinity in both cycles and increased as ambient salinity returned to the acclimation salinity. Total nitrogen excretion of stepwise acclimated sea cucumbers declined significantly from 30 to 15 S, but there was no salinity effect on total nitrogen excretion in sea urchins. Ammonia excretion varied directly with salinity in stepwise acclimated sea cucumbers (67–96% of total nitrogen excreted), but there was no salinity effect on ammonia excretion (89–95% of total nitrogen excreted) of sea urchins. Urea excretion did not vary with salinity in sea cucumbers (2–4% of total nitrogen excreted) or sea urchins (2–9% of total nitrogen excreted). Primary amines varied inversely with salinity in sea cucumbers (2–30% of total nitrogen excreted), but did not vary with salinity in sea urchins (2–4% of total nitrogen excreted). The oxygen: nitrogen ratio of both species indicated that carbohydrate and/or lipid form the primary catabolic substrate. The O:N ratio did not vary as a function of salinity. Both species are more tolerant to reduced salinity than previously reported, however, rates of oxygen consumption and/or nitrogen excretion are modified by salinity as well as season.     

Physiological responses of Nodularia harveyana to osmotic stress

The effects of salinity stress on biomass yield, photosynthetic O₂ evolution and nitrogenase activity were investigated using axenic cultures of Nodularia harveyana (Thwaites) Thuret originally isolated from a salt marsh at Gibraltar Point, Lincolnshire, UK in 1971 and studied in this laboratory in 1983. Biomass yields, as chlorophyll a per culture, were highest in the 0 to 100% seawater (0 to 35 sea salt) range with negligible growth in 200% seawater; growth on NH ₄ ⁺ was greater than on N₂ and NO ₃ ^- , which did not differ significantly from each other. In short-term experiments, photosynthetic O₂ evolution remained high at salinities up to 150% seawater (52.5 sea salt); nitrogenase activity remained high at salinities up to 100% seawater (35 sea salt). The major internal low molecular weight carbohydrate which accumulated in response to increased salinity was sucrose, the levels of which fluctuated markedly and rapidly in response to salinity change.     

Effects of salinity and bromine on zygotes and embryos of Fucus vesiculosus from the Baltic Sea

Zygotes and young embryos derived from Fucus vesiculosus collected in the archipelgo of Stockholm in 1990, growing at a salinity of 6 to 7 S, were cultured under different salinity conditions and in media of different bromine concentrations. Optimum salinity was 10 to 12 S for germination (rhizoid initiation) while apical hair formation showed a broader tolerance curve with an optimum at 8 to 14 S. Bromine caused inhibition of early development of F. vesiculosus. At 6 salinity a 50% reduction in germination took place at 10.0 mM Br and at 1.25 mM Br only 4.7% of the embryos developed apical hairs, as compared to 32.7% in the control. Bromine toxicity decreased at higher salinities. The results indicate that F. vesiculosus in the Baltic Sea has diverged from its Atlantic progenitors and to some extent acclimated to low salinity. Still, the salinity in the normal environment of the tested population is lower than optimum, leading to a lower degree of germination of zygotes, a lower growth rate of young embryos and probably also a higher sensitivity to additional stress factors such as chemical pollution.     

Salinity tolerance of benthic estuarine diatoms as tested with a rapid polarographic measurement of photosynthesis

Measurements of net photosynthesis of benthic estuarine diatoms were made by polarographic registration of oxygen saturation. A measuring cell was constructed in which media with salinities of 2.0 to 100.7 were pumped over the algae between measurements. Diatoms from unialgal cultures and mixed populations from intertidal flats appeared to be highly tolerant of extreme salinities. During short-term exposures (20 min) the net photosynthesis of the algae did not drop below 70% of the initial values, within the salinity range 4.0 to 60.0. Prolonged exposure (up to 6 h) gave essentially the same results. Populations of benthic diatoms, sampled from field stations with mean salinities of about 30, 12, and below 5, showed only gradual differences in their tolerance of salinities between 2.0 and 33.7. Two species, Navicula arenaria and Nitzschia sigma, were cultured in media ranging in salinity from 8.0 to 45.0 and from 2.0 to 45, respectively. The tolerance to changing salinity was only slightly affected by the salinity of the medium in the preculture. The role of salinity in the production and distribution of intertidal diatoms is discussed.     

Zooplankton of the Bristol Channel and Severn Estuary. The distribution of four copepods in relation to salinity

The seasonal variations in distribution and abundance of the common zooplankton species in the Bristol Channel and Severn Estuary were related to the salinity regimes observed over the period November 1973 to February 1975. The dominant constituents in all regions were the calanoid copepods, which reached maximum densities in July: approximately 100 times their winter levels. Four zooplankton assemblages were recognised using an objective classification program which computed similarity coefficients and used group-average sorting. The assemblages existed along the salinity gradient observed from the Severn Estuary to the Celtic Sea. The assemblages were classified as true estuarine, estuarine and marine, euryhaline marine and stenohaline marine and were characterized by the copepods Eurytemora affinis (Poppe) (<30S), Acartia bifilosa var. inermis (rose) (27 to 33.5S), Centropages hamatus (Lilljeborg) (31 to 35S) and Calanus helgolandicus (Claus) (>33S), respectively.     

Effects of salinity fluctuation on the hemolymph composition of the blue crab Callinectes sapidus

The hemolymph of the blue crab Callinectes sapidus was hyperosmotic during 20-10-20 S and 30-10-30 S diurnal cycles. The hemolymph became isosmotic at 26 S and hyposmotic at 28 S in the 10-30-10 S diurnal cycle. Hemolymph Na⁺ was hyperionic to seawater throughout all cycles. Hemolymph Cl^- was hyperionic below 24 S and either isionic or hypoionic from 24 to 30 S. Hemolymph K⁺ concentrations were hyperionic below 26 S and either isionic or hypoionic from 26 to 30 S. Hemolymph Mg⁺⁺ values were hypoionic over the experimental salinity range (10 to 30). Hemolymph ninhydrin-positive substances (NPS) levels were directly related to ambient salinity.