Heatstroke (HS)-induced acute lung injury (ALI) has high morbidity and mortality with no specific therapies. Ferroptosis, a form of programmed cell death driven by lipid peroxidation due to reduced Glutathione Peroxidase 4 (GPX4) activity, is closely linked to HS-induced ALI. This study investigated the effect of alpha-linolenic acid (ALA), a plant-derived ω-3 fatty acid, on ferroptosis in a mouse model of HS-induced ALI. Histopathology analysis found that ALA can attenuate lung injury and improve the 7-day survival rate in mice with HS-induced ALI. In addition, ALA significantly reduced the levels of reactive oxygen species (ROS) and malondialdehyde (MDA), while increasing the level of antioxidant glutathione (GSH). Further analysis showed that ALA upregulated the levels of SLC7A11 and GPX4 by promoting the nuclear translocation of Nrf2. This led to increased GSH synthesis but reduced ROS accumulation, which in turn suppressed ferroptosis and protected the mice against HS-induced ALI. Additionally, the protective effect of ALA was found to be diminished in Nrf2-deficient mice. In summary, ALA inhibits ferroptosis in macrophages by activating the Nrf2/SLC7A11/GPX4 pathway and attenuates HS-induced ALI.