The periodic application of chemical lampricides that selectively kill larval sea lampreys (Petromyzon marinus) in their nursery habitats remains a primary component of the Great Lakes Fishery Commission’s (GLFC) Sea Lamprey Control Program in the Laurentian Great Lakes. Lampricides include 3-trifluoromethyl-4-nitrophenol (TFM) and niclosamide, the 2-aminoethanol salt of 2′, 5-dichloro-4′-nitrosalicylanilide, which may be used as an additive to TFM during stream treatments, or alone in a granular, bottom-release formulation to target sea lamprey larvae in deepwater environments where dilution would render TFM ineffective. During the early 1990s, the GLFC identified lampricide reduction targets in response to societal concerns with pesticide use, rising lampricide costs, and promising research into alternative controls. By 1999, the GLFC’s control agents, Fisheries and Oceans Canada (DFO) and the U.S. Fish and Wildlife Service (USFWS), had reduced TFM use by 36%. However, without effective alternative methods to compensate for increasing larval and juvenile production, sea lamprey abundance and lake trout (Salvelinus namaycush) marking rates rose throughout the Great Lakes. Beginning in the early 2000s, the GLFC and its control agents responded to burgeoning sea lamprey populations by implementing measures to advance the use of lampricides, which included: 1) assessing and controlling sea lamprey larvae that survived treatment; 2) enhancing treatment efficacy; 3) developing new technology to effectively treat larval populations that inhabit deepwater environments; 4) increasing operational capacity to treat more tributaries and lentic areas at shorter intervals; and, 5) conducting large-scale and targeted treatment strategies. When comparing lampricide use between the decades of 1990–1999 and 2010–2019, significant increases occurred in the mean number of treatments and amounts of TFM and niclosamide applied annually. Concurrent with these actions, researchers undertook studies to identify factors that erode lampricide treatment efficiency, elucidate physiological mode of action, and investigate lethal and sub-lethal impacts of lampricide exposure on aquatic organisms. By integrating new operational tactics and strategies with advances in science and technology, the GLFC, DFO, and USFWS, with support from the U.S. Geological Survey and the U.S. Army Corps of Engineers, have achieved unprecedented suppression of sea lampreys and reduction in lake trout marking in the Great Lakes. However, emerging challenges potentially threaten the future use of lampricides.