In the aquatic environment, organisms are exposed to complex chemical mixtures throughout life, producing effects not anticipated in laboratory settings designed to test acute exposures of single chemicals. Exposure to chemical mixtures often produces results either not observed or counterintuitive to single chemical exposures. By employing field and laboratory-based exposures using fathead minnows and sunfish, exposures included sensitive life stages otherwise unobserved in adult acute exposure experiments. Field-based studies exposed fathead minnows and sunfish to water collected from sites along the Maumee River (Toledo, OH) to determine the impacts of a land use gradient (upstream – agriculture to downstream – industry and urban). Adult minnows were analyzed for fecundity, physiology, and hematological characteristics (vitellogenin, glucose, 11-keto testosterone, estradiol). Larval minnows were analyzed for growth, predator-avoidance behavior, feeding efficiency, and survival. Embryonic minnows were analyzed for viability, deformities, and time to hatch. Sunfish were either deployed at river sites or resident sunfish were collected for physiology, hematological characteristics (vitellogenin, glucose), and histological analysis. Results of these experiments demonstrate the effect of changing land use on aquatic organisms including reductions and delays in fecundity, alterations to metabolic indices, and greater severity of biological responses of fish exposed to waters from urban settings. Laboratory exposures analyzed the effects of eight co-occurring chemicals found in the Great Lakes watershed in areas of agricultural land use. Fathead minnows were exposed to a complex mixture of environmentally measured concentrations over three generations in a flow-through exposure system to assess the potential physiological, organism, and population level effects. Adult minnows were analyzed for physiology, hematological characteristics (vitellogenin, glucose), behavior, and fecundity. Larval fish were analyzed for growth, predator-avoidance behavior, and feeding efficiency. Juvenile fish were analyzed for growth. Adult minnows demonstrated reductions in fecundity at environmental concentrations in the second exposure generation. In addition, adults demonstrated increasing plasma vitellogenin and glucose, highlighting potential improper direction of energy. Larval and juvenile minnows were unaffected by mixture exposure. The alterations in both field and laboratory settings indicate the extent to which environmentally relevant agricultural mixtures have the potential to pose threats to aquatic organisms through reductions in fecundity, alterations to plasma proteins, and changes in physiology associated with contaminant exposure. Further, the use of a laboratory study demonstrates the need for complete life-cycle assessment of contaminants as indicated by differences between first and second-generation responses. Agricultural practices, and associated aquatic pollution, pose a threat to both the organism and population level of two North American species.