Bioavailability of metals to fish: its molecular basis, toxicological consequences and how it can be assessed Christer Hogstrand Metal Metabolism Group King s College London Global maps for the year 2000 of in-use (A) copper stock intensities (Gg/grid cell) compared with in-ground (B) copper resources Rauch J N PNAS 2009;106:18920-18925
Relationship between the background concentration of zinc in water and toxicity expressed as the no observed effect concentration (NOEC) Bodar et al., (2005) Integr Environ Assess Manag 1 Metal concentrations in The River Hayle, Cornwall
Comparative acute toxicity of metals to rainbow trout McDonald (1982) The teleost gill is a target organ for metal toxicity Eckert et al. (1997) Animal Physiology
Na + Water Gill Plasma 0.1 20 mosmol 300 mosmol Na + 3 Na + ATP H + Mg ATP 2 K + NH 3 H + NH 3 Na + H +, NH 4 Zn CA CO 2 HCO 3 - K + Cl - Ca 2+ HCO 3-, OH - GSH MT Cl - Ca2+ ATP? H 2 O O 2 Apical -70 mv Basolateral CO 2 Ca 2+ Cl -
Zinc inhibits calcium uptake leading to acute hypocalcaemia Am. J. Physiol. 270 Speciation dictates metal bioavailability and toxicity http://www.lab-initio.com/screen_res/nz015.jpg
Metal impact on biota in Big Bayou Creek, Kentucky Birge et al. (2000) Environ. Toxicol. Chem. 19 Impact correlates to bioavailable rather than total dissolved metal Calculated bioavailable Cu, Ag and Cd fractions correlated significantly with BA score (r > -0.95) and number of taxa ( r = -0.95). Birge et al. (2000) Environ. Toxicol. Chem. 19
Fish gill physiology
Expression of zinc transporters in gill changes in response to changes in water zinc concentrations Zheng et al. (2008) Physiol Genomics 34: 205 214,.
Physiology modifies metal affinity and accumulation Walker et al. Environ. Sci. Technol. 2007, 41, 6505-6513; Toxicol. Appl. Pharmacol. 2008, 230, 67 77
Fish Gill In vitro Cell culture System (FIGCS) is a functional transporting epithelium TER KΩ cm -1 [Zn 2+ ] Walker et al. Environ. Sci. Technol., 2007, 41, 6505-6513 Comparison of effects of waterborne Ag(I) on rainbow trout and FIGCS Walker et al 2008 Toxicol Appl Pharmacol 230: 67-77
Comparison of the effects of water chemistry on whole organism ion uptake and gene expression response in vitro ZnT1 MTs Walker et al., (2008) Toxicol. Appl. Pharmacol. 230(1): 67-77 Chronic zinc toxicity Species Average NOEC (µg/l) Jornanella floridae 44 2 Eurasian minnow 50 1 Fathead minnow 78? Rainbow trout 189 1 Brook trout 530? Zebrafish 660 9 Number of studies Fish toxicity data used in the EU Environmental Risk Assessment for Zn End-points to set NOEC were: -Survival -Reproduction -Growth Species LOEC Response Hardness ph Reference µg/l mg/l CaCO3 Rainbow trout 5.6 Avoidance 14 7.2 Sprague (1968) Lake whitefish 10 Avoidance 90 7.6 Scherer and McNicol (1998) Rainbow trout 10 Avoidance 248 8 Svecevicius (1999) Rainbow trout 47 Avoidance 112 7.6 Black and Birge (1980) Atlantic salmon 53 Avoidance 18 7.5 Sprague (1964) Rainbow trout 144 Ventilation rate 25 7 Cairns et al. (1982) Vimba bream 220 Avoidance* 120 7.3 Svecevicius (1999) Brook charr 1,390 Cough rate 45 7.5 Drummond and Carlson (1977) Bluegill 3,640 Movement pattern 51 7.8 Waller and Cairns (1972)
Normal Water: 0.5 um Diet: 1.2 nmol g -1 day -1 Excess Water: 5 um Diet: 120 nmol g -1 day -1 n=9 Zheng et al. BMC Genomics 2010, 11:553 Annotation Enrichment on day Clusters Category Name Genes 0.3 1 4 7 14 P-value Development BP developmental process 96 0.7 0 0 0 0.1 7.4E-06 BP anatomical structure development 68 0.4 0 0.1 0 13.6E-05 BP system development 54 0.2 0.2 0.2 0 16.2E-04 BP multicellular organismal process 97 0.5 0.4 0 0 0.6 1.1E-03 BP nervous system development 29 0.3 0.5 0 0 11.2E-03 BP cellular developmental process 53 0.8 1 1 0 0.5 2.7E-03 BP cell differentiation 53 0.8 1 1 0 0.5 2.7E-03 BP cell development 39 0.5 1 1 1 0.2 3.1E-03 BP multicellular organismal development 64 0.4 0.4 0.2 0 13.4E-03 BP brain development 10 1 0.1 1 1 1 3.9E-03 BP central nervous system development 13 1 0.2 0.1 1 1 4.7E-03 BP organ development 39 0.3 0.1 0.4 0.1 1 5.0E-03 BP cell fate commitment 7 1 1 1 1 1 7.4E-03 Metabolism BP primary metabolic process 197 0.1 0 1 0 0.3 1.1E-05 BP metabolic process 213 0.1 0 1 0 0.1 1.2E-05 BP cellular metabolic process 196 0 0 1 0 0.1 1.5E-05 BP lipid biosynthetic process 14 1 0.2 1 1 1 3.0E-03 BP biopolymer metabolic process 126 0.1 0 1 0.2 0.6 4.9E-03 BP macromolecule metabolic process 161 0.5 0 1 0.1 0.6 8.2E-03 BP multicellular organismal catabolic process 4 1 1 1 1 1 8.3E-03 Gene expression MF sequence-specific DNA binding 28 1 0 1 0 17.6E-06 BP transcription from RNA polymerase II promoter 28 0.3 0 1 0.1 12.4E-04 BP positive regulation of nucleobase, nucleoside, nucleotide and nucleic acid metabolic process 16 1 0 1 1 1 2.3E-03 MF transcription factor activity 33 1 0 1 0 13.0E-03 BP positive regulation of transcription 15 1 0 1 1 1 4.4E-03 BP positive regulation of transcription, DNA-dependent 13 1 0 1 1 1 5.1E-03 MF transcription factor binding 17 1 0 1 1 1 5.3E-03 Nuclear receptors BP response to steroid hormone stimulus 6 1 1 1 0 16.8E-04 MF steroid hormone receptor activity 6 1 1 1 1 1 5.0E-03 MF retinol binding 3 1 1 1 1 1 8.9E-03 MF ligand-dependent nuclear receptor activity 6 1 1 1 1 1 9.0E-03 Regulation BP positive regulation of biological process 38 0.2 0 1 0 17.2E-04 BP positive regulation of cellular process 33 1 0 1 1 1 3.2E-03 BP regulation of multicellular organismal process 14 1 1 0.2 0 14.5E-03 BP regulation of biological process 114 0.6 0.2 1 0 0.8 4.7E-03 BP biological regulation 122 0.5 0.1 1 0 0.9 8.3E-03 Others MF oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen 9 1 1 0 0.2 1 3.0E-03 BP microtubule cytoskeleton organization and biogenesis 7 1 1 1 0.1 15.3E-03 BP cell cycle process 26 1 1 0.1 0 15.5E-03 MF 5'-nucleotidase activity 3 1 1 0 1 1 6.7E-03 MF oxygen binding 5 1 1 1 1 1 6.8E-03 MF iron ion binding 14 1 1 0.1 0.1 17.3E-03 MF nucleotidase activity 3 1 1 0 1 1 8.9E-03 BP detection of abiotic stimulus 5 1 1 1 1 1 9.2E-03 >0.1 <0.1 <0.05 <0.01 <0.005 <0.001 Development Metabolism Gene expression Nuclear receptors Regulation Others (e.g. Cell cycle process, iron binding, microtuble cytoskeleton organisation) Zheng et al. BMC Genomics 2010, 11:553
Zn 2+ is a potent inhibitor of protein tyrosine phosphatases RPTP-β, K i = 21 ± 7 pm 2012 by American Society for Biochemistry and Molecular Biology Wilson et al. (2012) J Biol Chem 287, 9322 Zn 2+ inhibits enzymes at physiological [Zn 2+ ] i 1 0 Saturation of Zn binding sites in proteins ( ) 0.5 0 [Zn 2+ ] i 6 8 10 12 14 0.5 1 Saturation of Zn binding sites in MT ( ) pzn ( log[zn 2+ ]) Modified from Maret (2011) J Biol Inorg Chem 16:1079 1086
Total cumulative number of eggs 7000 6000 5000 4000 3000 2000 1000 No. of eggs Total cumulative number of spawns 60 50 40 30 20 10 No. of spawns 0 40 45 50 55 60 65 70 0 40 45 50 55 60 65 70 Day Day Metal Concentration (μg g -1 dry weight) 800 600 400 200 20 15 10 5 0 * * * * * Ag As Cd Cu Fe Zn Mean hatch rate (%) 120 100 * 150 * Mean number eggs per spawning pair 80 120 60 90 40 60 20 30 0 0 35 40 45 50 55 60 65 70 75 35 40 45 50 55 60 65 70 75 Day Day 2.5 2.0 Hatch rate Gene express 1.5 1.0 0.5 0.0 Vtg MT-II 180 Eggs/spawn Environ. Sci. Technol. 2008, 42, 5354 5360 Metal storage strategies in Gammarus Khan et al., Aquat. Toxicol. 96 (2010) 124 129 / J. exp. Biol., in revision
Effect of metal storage strategy on dietary metal uptake and toxicity in zebrafish Khan et al., 2009 Aquat. Toxicol. / 2010 Aquatic Toxicol Acknowledgments King s Dr Nic Bury Dr Wolfgang Maret Dr Dongling Zheng Mr Phil Cunningham Dr Andong Qiu Dr David Boyle Dr Paul Walker Dr Farhan Khan Dr Issa Muraina Mr Matt Wilson Dr Peter Kille Dr Graham Feeney Prof Richard Handy Dr Rod Wilson Dr Fernando Galvez Prof Chris Wood Prof Wesley Birge Mr Steve Munger Dr Rick Playle