Publication Date

2009-08-06

Availability

Open access

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PHD)

Department

Marine Biology and Fisheries (Marine)

Date of Defense

2009-06-12

First Committee Member

Martin Grosell - Committee Chair

Second Committee Member

Patrick Walsh - Committee Member

Third Committee Member

Chris Wood - Committee Member

Fourth Committee Member

Douglas Crawford - Committee Member

Fifth Committee Member

Frank Millero - Outside Committee Member

Abstract

Our knowledge of aquatic Cu toxicity has increased greatly over the past several years culminating with the incorporation of a model (the Biotic Ligand Model (BLM)) into the regulatory framework which allows for the site specific adjustment of water quality criteria based on water chemistry. However, our understanding of Cu toxicity in the aquatic environment is limited mostly to freshwater (FW). Because of this limited knowledge, this dissertation set out to examine the affect of salinity on Cu toxicity and accumulation across salinities from FW to sea water (SW). First, this work examined tissue specific Cu accumulation in five salinities (FW, 5 ppt, 11 ppt, 20 ppt, 28 ppt) from waterborne Cu exposure at two [Cu] (30 and 150 µg Cu L-1) in the euryhaline killifish, Fundulus heteroclitus. Branchial and hepatic accumulation followed a pattern that would be expected based on speciation and competition from cations. [Cu] were high in FW and decreased as salinity increased. However, in the intestine, [Cu] were highest at 5 ppt and were also elevated in the higher salinities. The elevation at the higher salinities was most likely due to drinking by the fish which increases as salinity increases above the isoosmotic point of the fish (~10 ppt) for osmoregulatory purposes and showed a trend toward increasing [Cu] with increasing salinity as would be expected. Secondly, the mechanism of Cu toxicity in FW and SW was examined in killifish. The mechanism of Cu toxicity in the killifish in FW was the same as had been seen for other FW fish. Cu exposure caused a decrease in Na+ / K+ ATPase activity which led to a decrease in whole body [Na+] which is the likely cause of death. In SW, surprisingly no ionoregulatory disturbances were observed. The only measured parameter that was changed in SW was net ammonia which showed a substantial decrease. Therefore, the mechanism of acute copper toxicity in FW and SW differed suggesting that physiology may need to be considered in future development of a BLM for SW. Next, the effect of salinity on Cu accumulation from a naturally incorporated dietary source was examined in FW and SW in Fundulus heteroclitus. Cu accumulation was not seen to differ in the two salinities in spite of differences in gut fluid chemistry that would lead to an ~11 fold difference in free Cu ion between FW and SW. This indicated that Cu accumulation from a dietary source was not as the free ion but most likely as organically (amino acid) bound Cu. In addition to this, subcellular fractionation of the diet revealed that Cu available to the next trophic level did not change with increasing waterborne [Cu], increasing tissue [Cu], or time in oysters which has implications for Cu accumulation in the environment. Finally, the molecular mechanisms behind effects and acclimation to waterborne and dietary Cu exposure in FW and SW were examined using suppression subtraction hybridization in the killifish intestine. Exposure to Cu induced a stress response which could be responsible for upregulation of genes involved in protein synthesis, proteolysis and ATP production. At the tissue level, two main responses were observed. First, genes necessary for muscle function were upregulated potentially in response to previously observed decreased intestinal motility. Secondly, apoptotic genes were upregulated corresponding to increased rates of apoptosis in intestines during dietary exposures. At the cellular level, metal chelators and oxidative stress genes were upregulated in response to increased free Cu in the cell and the subsequent free metal induce free radical formation. This work has advanced our understanding of Cu toxicology in saline environments and emphasized the importance of considering both chemistry and physiology in analyzing and interpreting Cu toxicology especially in saline environments. Hopefully, it will contribute to the future development of Cu water quality criteria.

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