Environ. Sci. Technol. 2001, 35, 669-675
A Mechanistic Model for the Uptake of Waterborne Strontium in the Common Carp (Cyprinus carpio L.) MOHAMMED J. CHOWDHURY AND RONNY BLUST* Department of Biology, University of Antwerp (RUCA), Groenenborgerlaan 171, B-2020 Antwerp, Belgium
The release of radioactive strontium to the environment is of concern due to the strong accumulation of this calcium resembling element in the bone and other tissues. To predict the effects of changes in environmental conditions on the uptake of Sr2+ and Ca2+ by freshwater fish, a Michaelis-Menten type model is introduced that accounts for the effects of chemical speciation, hydrogen ion activity, and metal ion competition. The uptake kinetics were characterized in vivo from short-term exposure experiments using the common carp (Cyprinus carpio) as the model organism. Fish were exposed to a wide range of waterborne Sr2+ (0.2-10 000 µM) and Ca2+ (1010 000 µM) concentrations and water pH (5.0-8.5). Strontium uptake by the whole body of fish increased with increasing Sr2+ activity, displaying saturation kinetics, but decreased significantly with increasing Ca2+ and H+ activities in the water. Likewise, calcium uptake by the fish decreased with increasing Sr2+ and H+ activities in the water. The model fitted to the pooled data explains 97.5% of the variation in Sr2+ uptake and 86% in Ca2+ uptake over the wide range of exposure conditions and reveals that Sr2+ and Ca2+ inhibit each other completely competitively, while H+ inhibits the uptake of both metal ions in a partially noncompetitive way. This model can be used as a mechanistic tool to predict the uptake of these metals in carp under variable conditions.
embedded systems (e.g., channels, carriers, or pumps) that transport the metal across the gill epithelium. The gating properties of these systems depend on the environmental conditions such as the hydrogen ion activity or the presence of other metal ions (12, 13). So far a mechanistically based model for Sr2+ uptake in freshwater fish has not been developed. Such a model takes the effects of water chemistry and the processes involved in metal transport across the membrane interface into account in a rational way. Recently, we have shown that Sr2+ and Ca2+ interact competitively for the same uptake site in the gills of carp and that the effect of waterborne Ca2+ on Sr2+ uptake and vice versa can be accurately described by a Michaelis-Menten type competitive inhibition model (8). Another important environmental factor is the hydrogen ion activity (pH) of the water. Water pH can influence Sr2+ uptake in three distinct ways: (i) changing the speciation of the metal in the solution; (ii) modulating the activity of the transport system; and (iii) affecting physiological processes that either directly or indirectly influence the uptake process (e.g., altering membrane potentials, ionic, and osmotic regulation) (12-14). The effect of changes in pH on metal uptake and toxicity is well-documented, but the types of interaction involved are poorly characterized (12, 15, 16). This information is required for the development of a mechanistic model for the effect of changes in environmental conditions on metal uptake. Recently, biotic ligand models have been introduced to account for the effects of chemical speciation and some other environmental factors on metal uptake and toxicity. These models assume that cations, such as proton and calcium, compete for the same binding sites on the cell surface (17-19). To understand the combined effect of chemical and biological processes on Sr2+ uptake in fish, we acclimated small carp (3-8 g) to different pH and determined the effect of pH on uptake from short-term exposure experiments. Our central goal was to develop a mechanistic model that is based on chemical and biological principles and can predict Sr2+ uptake in carp under different strontium, calcium, and pH scenarios. Additionally, we report here that H+ inhibits Sr2+ and Ca2+ uptake in a partially noncompetitive manner.
Materials and Methods Introduction The release of strontium to the environment as its radionuclides (90Sr and 89Sr) is an important environmental problem because of the calcium-like behavior of the element in biological systems resulting in the strong accumulation of the radionuclides in calcareous tissues (1, 2). The major sources of the radionuclides in aquatic environments are nuclear installations, dumping and leakage of radioactive wastes, and accidental release (3). The uptake of strontium (Sr2+) by aquatic organisms strongly depends on the environmental conditions and the functional characteristics of the exchange structures involved in solute exchange. The concentration of waterborne calcium is the most important variable controlling the uptake of Sr2+ (4-8), and empirical models to predict the concentration factor of strontium by freshwater fish were developed (6, 9). It has been reported that Sr2+ uptake occurs mainly via the gills and that uptake via food is of secondary importance (10, 11). Strontium uptake is facilitated by membrane * Corresponding author telephone: 32-3-2180347; fax: 32-32180497; e-mail:
[email protected]. 10.1021/es000142t CCC: $20.00 Published on Web 01/10/2001
2001 American Chemical Society
Experimental Fish and Medium. Fingerlings (∼1.5 g) of the common carp, Cyprinus carpio, were obtained from the fish hatchery of the Agricultural University of Wageningen, The Netherlands. They were grown for at least 3 months in the laboratory at 25 ( 1 °C and pH 7.6-8.0 following a standard protocol for water quality and feeding (8). Unless otherwise stated, the acclimation and exposure medium for all experiments was standard medium-hard freshwater prepared by dissolving the reagent-grade salts (CaCl2, 348; MgSO4, 500; KCl, 54; and NaHCO3, 1143 µM) in deionized water (20). Stable strontium was added to the exposure medium as the SrCl2 salt. Experimental Procedures. Two experiments were conducted to determine the effect of pH acclimation and exposure on Sr2+ and Ca2+ uptake. Using these experimental results together with existing data, a model was constructed for the effect of pH and Ca2+ on Sr2+ uptake in carp. Since acclimation to pH can change the membrane characteristics and physiology of fish, we started our study with an acclimation experiment. In this experiment, fish were acclimated to different pH in four acclimation groups (pHaccl ) 5, 6, 7, and 8) at 25 ( 1 °C for 16 d. After being acclimated, each group was exposed to different pH in five exposure VOL. 35, NO. 4, 2001 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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groups (pHexp ) 5, 6, 7, 8, and 9) at 25 ( 0.5 °C for 3 h creating different combinations of acclimation and exposure. During acclimation, 35-40 individuals were held in a polyethylene tank containing 150 L of standard water. The pH was controlled by a pH-stat system (Consort, Belgium) that kept the pH within narrow limits by adding HCl or NaOH with a drifting value of 0.90, p < 0.001, n ) 98) and Ca2+ (r > 0.80, p < 0.001, n ) 100). The observed increase in Sr2+ and Ca2+ uptake with increasing pH of exposure within each acclimation group is consistent with results for some other metals such as copper, cadmium, and zinc (14, 16, 28). Competition between the metal ions and protons for binding sites in the transport system has been proposed as the reason for this effect (19). The significant acclimation effect indicates that the transport characteristics of the Sr2+/Ca2+ uptake system changed during VOL. 35, NO. 4, 2001 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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TABLE 1. Two-way Analysis of Variance Contrasting Effects of pH of Acclimation,a pH of Exposure,b and Their Interactionc on the Uptake of Sr2+ (Bold Type) and Ca2+ (Normal Type) in Whole Body, Gills, and Blood of Carpd source of variation (effect) I II I × II
I
FIGURE 1. Uptake of Sr2+ in the whole body (circles), gills (triangles), and blood (squares) of carp as a function of exposure pH for the different groups of acclimation pH (pHaccl). Data points represent mean uptake with standard deviations in 3 h (n ) 5-6; Srtotal ) 0.21 µM; Catotal ) 348 µM; temperature ) 25 ( 0.5 °C). Open symbols are the data points obtained when acclimation pH and exposure pH were the same.
II I × II
I II I × II
dfe effect
MSf effect
3 3 4 4 12 12
2.36 3.24 3.51 7.71 0.24 0.28
3 3 4 4 12 12
1.70 2.06 5.00 17.13 0.24 0.40
Gill 79 81 79 81 79 81
3 3 4 4 12 12
1.68 4.78 3.92 4.83 0.20 0.14
Blood 79 81 79 81 79 81
df error
MS error
F
p
40.46 51.65 60.22 122.98 4.11 4.43
