Aquatic Toxicology 65 (2003) 377–395 Antioxidant defenses in killifish (Fundulus heteroclitus) exposed to contaminated sediments and model prooxidants: short-term and heritable responses Joel N. Meyer a , Jim D. Smith a , Gary W. Winston b , Richard T. Di Giulio a,∗ a Nicholas School of the Environment and Earth Sciences and Integrated Toxicology Program, Duke University, Durham, NC 27708-0328, USA b Department of Environmental Health, Ministry of Health, Jerusalem 91010, Israel Received 4 October 2002; received in revised form 30 May 2003; accepted 20 June 2003 Abstract A population of killifish (Fundulus heteroclitus) inhabiting a Superfund site on the Elizabeth River (VA, USA) is tolerant of the acute toxicity of the sediments from the site; previous work suggests that this tolerance is based both on genetic adaptation and physiological acclimation. In this study, larval first- and second-generation (F 1 and F 2 ) offspring of Elizabeth River killifish were more resistant to the toxicity of t-butyl hydroperoxide (a model prooxidant) than were King’s Creek (reference site) offspring, indicating a heritable tolerance of exposure to oxidative stress. In laboratory experiments designed to elucidate the mechanistic basis for this increased tolerance, we exposed laboratory-raised F 1 and F 2 offspring from Elizabeth River and King’s Creek killifish to Elizabeth River sediments, menadione, or t-butyl hydroperoxide, and measured the following antioxidant parameters: total oxyradical scavenging capacity (TOSC); glutathione content (total and disulfide); activities of glutathione reductase (GR); glutathione peroxidase (GPx); and glutamate cysteine ligase (GCL) activities and protein levels of copper–zinc superoxide dismutase (CuZnSOD); and protein levels of manganese superoxide dismutase (MnSOD). Exposure to Elizabeth River sediments lead to consistent increases in total glutathione concentrations, GR activities, and MnSOD protein levels, and in some cases increased GPx and GCL activities, in both populations. In addition, Elizabeth River offspring (larvae) showed higher basal TOSC values, glutathione concentrations, and MnSOD protein levels. These data suggest that upregulated antioxidant defenses play a role in both short-term (physiological) and heritable (multigenerational/evolutionary) tolerance of the toxicity of these Superfund sediments. The responses of specific antioxidant parameters, including sex-specific responses in the cases of glutathione concentrations and GR activity, are also discussed. © 2003 Elsevier B.V. All rights reserved. Keywords: Oxidative stress; Polycyclic aromatic hydrocarbons; Adaptation; Superfund; Creosote; Fundulus heteroclitus ∗ Corresponding author. Tel.: +1-919-613-8024; fax: +1-919-684-8741. E-mail address: richd@duke.edu (R.T. Di Giulio). 1. Introduction Atlantic killifish (Fundulus heteroclitus; also known as mummichog) inhabiting a contaminated site on the Elizabeth River (Norfolk, VA) are tolerant of the 0166-445X/$ – see front matter © 2003 Elsevier B.V. All rights reserved. doi:10.1016/j.aquatox.2003.06.001