Pesticide use has increased significantly in recent decades. While pesticides play a crucial role in food production, their widespread application raises serious environmental concerns. Most research to date has focused on lowland regions, yet recent evidence suggests that mountain ecosystems are also at risk due to both local agricultural practices and the long-range atmospheric transport of pesticides. This study investigates pesticide contamination and ecological risks in five mountainous agricultural watersheds in Chile—Limarí, Choapa, Aconcagua, Maipo, and Cachapoal—where pesticide use supports intensive crop production. Using passive samplers (POCIS) and sediment analysis, we found widespread pesticide contamination, with detections at 26 of 30 sites, varying notably across locations. A total of 20 compounds were identified, with different detection profiles depending on the sampling method: herbicides, primarily DEA, were more prevalent in POCIS, whereas insecticides dominated in sediments. The highest contamination levels occurred in the southernmost Maipo and Rapel watersheds, with up to eight pesticides detected in certain locations. Water conductivity, agricultural land cover, and latitude were identified as key factors influencing pesticide distribution. Ecotoxicological risk assessments revealed that eight pesticides exceeded high-risk thresholds for aquatic organisms, particularly insecticides and fungicides. Pyrethroids—deltamethrin, cyfluthrin, and lambda-cyhalothrin—posed severe threats to fish and invertebrates. High-risk pesticide levels were also detected in sediments, especially in the Limarí watershed. These findings highlight the urgent need for enhanced pesticide monitoring and regulation in mountain freshwater ecosystems of Chile, which provide essential water resources and support unique biodiversity. This research represents one of the first comprehensive assessments of pesticide risks in mountainous rivers, underscoring the ecological threats posed by agricultural contaminants.