Cultural eutrophication—the process by which human activities increase nutrient input rates to aquatic ecosystems and thereby cause undesirable changes in surface-water quality—is generally thought to have begun with the start of the industrial era. The prehistoric dimension of human impacts on aquatic ecosystems remains relatively undescribed, particularly in North America. Here we present fossil plankton data (diatoms and rotifers), organic and inorganic carbon accumulations, and carbon isotope ratios from a 1000 yr sediment core record from Crawford Lake, Ontario, Canada. The data document increased nutrient input to Crawford Lake caused by Iroquoian horticultural activity from A.D. 1268 to 1486 and show how this increased nutrient input elevated lake productivity, caused bottom-water anoxia, and irreversibly altered diatom community structure within just a few years. Iroquoian settlement in the region declined in the fifteenth century, yet diatom communities and lake circulation never recovered to the predisturbance state. A second phase of cultural eutrophication starting in A.D. 1867, initiated by Canadian agricultural disturbance, increased lake productivity but had comparatively less impact on diatom assemblages and carbon-storage pathways than the initial Iroquoian disturbance. This study deepens our understanding of the impact of cultural eutrophication on lake systems, highlights the lasting influence of initial environmental perturbation, and contributes to the debate on the ecological impacts of density and agricultural practices of native North American inhabitants.