Native habitats have been altered by human activity to create spaces for human use, such as urbanization and agriculture. These alterations often disrupt the natural landscape, resulting in habitat isolation, loss, and fragmentation. Creeks can be influenced by human development, and such development can impact many creek-dwelling species. Drastic changes in pH or dissolved oxygen can disrupt the creek biota, including alteration of fish assemblages and health. The purpose of this study was to determine if different riparian land alterations affect the water quality of various creeks and the physiology of creek-dwelling fish, using Redfin Shiners as a model species. Redfin Shiners were captured with a seining net at 12 different sites among 4 different creeks in Central Illinois. These sites could generally be classified as residential, agricultural, or riparian areas protected from development. We also recorded a suite of water chemistry data. We captured 271 Redfin Shiners among the sites, placing each into hormone-free spring water for 30 minutes, allowing them to exude cortisol and then measuring cortisol extracted from water samples. We found higher cortisol levels among fish in creeks with lower average dissolved oxygen over the 8-week study. Creek sites immediately downstream from agricultural areas had the lowest dissolved oxygen and shiners with the highest cortisol levels; whereas fish in the areas protected from development had the lowest cortisol levels. Our findings demonstrate that significant alteration of the stream habitat and associated changes in stream chemistry can impact glucocorticoid levels in native, pool-dwelling fish.