Grantee Research Project Results
2002 Progress Report: Behavioral Toxicology and Pathology of Early Exposure to Toxic PCOs
EPA Grant Number: R828224Title: Behavioral Toxicology and Pathology of Early Exposure to Toxic PCOs
Investigators: Kane, Andrew S.
Current Investigators: Kane, Andrew S. , Salierno, James D.
Institution: University of Maryland - College Park
Current Institution: University of Maryland - College Park , University of Maryland School of Medicine
EPA Project Officer: Packard, Benjamin H
Project Period: September 1, 2000 through August 30, 2003 (Extended to April 30, 2005)
Project Period Covered by this Report: September 1, 2001 through August 30, 2002
Project Amount: $423,264
RFA: Ecology and Oceanography of Harmful Algal Blooms (1999) RFA Text | Recipients Lists
Research Category: Water , Aquatic Ecosystems
Objective:
The overall objective of this research project is to detect and differentiate exposure to harmful algal bloom (HAB) stressors using integrated, whole-organism behavioral and tissue-level responses in fish. Mummichog (Fundulus heteroclitus) is a widely distributed, nonmigratory euryhaline fish species that thrives in many brackish waters in the mid-Atlantic United States, is an ecologically important species, and is readily maintained in the laboratory. Our primary objective is to develop the mummichog model to detect low-level HAB stress using behavioral toxicologic methods. Tilapia and bluegill (Lepomis macrochirus) also were investigated in different portions of this research project. Furthermore, we are evaluating immediate/early responses in the brains of exposed fish by quantitatively detecting alterations in energy utilization or gene expression.
Progress Summary:
Behavioral Toxicology. Year 1 of the project was dedicated to validating the new behavioral toxicology system, and initial datasets indicated that mummichog is a good model to investigate behavioral effects of HAB exposure. We validated our exposure system with respect to accurately tracking fish movement paths through various acclimation experiments and exposure to a model anesthetic drug, MS-222. Exposure to a light anesthetic dose for mummichog, 60 mg/L, determined through preliminary EC50 exposures, was associated with statistically significant alterations in the following endpoints: percent movement, velocity, fractal dimension, and distance from center and startle response. Once we gained confidence in our ability to quantitatively and repeatedly track the movement of individual fish and detect differences between groups, we initiated work with brevetoxin, PbTx, a model HAB toxin. Brevetoxin is one of the most significant HAB toxins worldwide with respect to both commonality of occurrence and ecological effects. Following dose-response experiments to determine the LC50 and EC 50 for PbTx, we exposed mummichog to PbTx-2 at 40 µg/L, a concentration that is "low to moderate" with respect to environmental relevancy. Data from these exposures indicated that PbTx-2 has subtle effects on mummichog movement and behavior, with significant alteration only in the startle response endpoint. Fish were neither attracted to PbTx-2, nor did they avoid it, based on space utilization data. At this low concentration, there were no statistically relevant changes regarding velocity, angular change, burst swimming, or path complexity (i.e., fractal dimension) endpoints.
Because the objective of applying behavioral endpoints was to qualitatively and quantitatively describe varying responses to different HAB exposures, in Year 2 of the project, we modified and expanded our analytical software capabilities to discern movement changes as well as group dynamics with exposures to multiple fish in each exposure arena. Validation with this new software is now complete and we will implement sublethal exposures using groups of fish to: PbTx-2 (higher, but still environmentally relevant concentrations), saxitoxin (STx), and Cheatocerus. Behavioral observations of Pfiesteria piscicida-exposed Tilapia also were conducted at North Carolina State University (NCSU) in collaboration with JoAnn Burkholder, Howard Glasgow, Nora Deamer-Melia, and Mathew Parrow. Multiple groups of tilapia were exposed to active P. piscicida cultures in glass aquaria, and their behavior was videographically recorded. Initial analyses of these data indicate that there were notable alterations in both social interactions as well as behavioral patterns associated with exposure.
Neurobiology Studies. In collaboration with Ellen Silbergeld, Jennifer Choich, and James Salierno, L. macrochirus were exposed to a low, environmentally relevant PbTx concentration (or control water) and injected with radiolabeled 2-deoxyglucose. PbTx-exposed fish have significantly higher midbrain labeling as compared with control fish. However, the labeling is relatively diffuse. Subsequent efforts with mummichog focused on applying c-fos detection methods in an attempt to gain better regional specificity. Mummichog were exposed to brevetoxin, domoic acid, transport stress, and in collaboration with Dr. Jeffrey Shields, to live cultures of Pfiesteria shumwayae. Brains of exposed fish were subsequently processed for changes in c-fos expression. Preliminary data suggest that there are regional differences in this immediate/early-response gene for certain exposure types.
Future Activities:
We will focus our efforts in the remaining years of this project on completing the behavioral toxicology studies with PbTx, STx and Cheatocerus in groups of mummichog, and enumerating changes in c-fos expression in exposed mummichog brains. These studies will differentiate early, subtle, quantifiable changes in behavior and movement associated with exposure to low-level HAB stressors. These changes may be linked to possible changes occurring in the central nervous system of exposed animals. Additional activities include the development of an outreach Web site through the University of Maryland Aquatic Pathobiology Laboratory.
Journal Articles:
No journal articles submitted with this report: View all 9 publications for this projectSupplemental Keywords:
c-fos, 2-deoxyglucose, mummichog, bluegill, tilapia, behavioral toxicology, fish pathology, ulcerative lesions, harmful algal blooms, video motion analysis, Pfiesteria, brevetoxin, saxitoxin, domoic acid, outreach., RFA, Scientific Discipline, Geographic Area, Waste, Water, Ecosystem Protection/Environmental Exposure & Risk, Limnology, Contaminated Sediments, exploratory research environmental biology, Ecosystem/Assessment/Indicators, Chemical Mixtures - Environmental Exposure & Risk, Ecosystem Protection, Chemistry, State, Ecological Effects - Environmental Exposure & Risk, algal blooms, Ecological Effects - Human Health, Southeast, Biology, East Coast, Ecological Indicators, ecological exposure, pfiesteria piscicida, North Carolina, agricultural runoff, ecology, bloom dynamics, fish kills, Shephard's Crook, whole-fish bioassay, suburban watersheds, economic assessments, fish lesions, Gyrodiniums, harmful algal blooms, ligand sensitive hosts, Everglades, environmental chemistry, pfiesteria, toxins, bioassay, dinoflagellate, Glenodriniums, Tilapia, Florida, fluorescence sensing, Virginia, ScripsiellasProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.