Climate change is causing large declines in many Pacific salmon populations. In particular, warm rivers are associated with high levels of premature mortality in migrating adults. The Fraser River watershed in British Columbia, Canada, supports some of the largest Chinook salmon (Oncorhynchus tshawytscha) runs in the world. However, the Fraser River is warming at a rate that threatens these populations at critical freshwater life stages. A growing body of literature suggests salmonids are locally adapted to their thermal migratory experience, and thus, population-specific thermal performance information can aid in management decisions. We compared the thermal performance of pre-spawning adult Chinook salmon from two populations, a coastal fall-run from the Chilliwack River (125 km cooler migration) and an interior summer-run from the Shuswap River (565 km warmer migration). We acutely exposed fish to temperatures reflecting current (12°C, 18°C) and future projected temperatures (21°C, 24°C) in the Fraser River and assessed survival, aerobic capacity (resting and maximum metabolic rates, absolute aerobic scope (AAS), muscle and ventricle citrate synthase), anaerobic capacity (muscle and ventricle lactate dehydrogenase) and recovery capacity (post-exercise metabolism, blood physiology, tissue lactate). Chilliwack Chinook salmon performed worse at high temperatures, indicated by elevated mortality, reduced breadth in AAS, enhanced plasma lactate and potassium levels and elevated tissue lactate concentrations compared with Shuswap Chinook salmon. At water temperatures exceeding the upper pejus temperatures (Tpejus, defined here as 80% of maximum AAS) of Chilliwack (18.7°C) and Shuswap (20.2°C) Chinook salmon populations, physiological performance will decline and affect migration and survival to spawn. Our results reveal population differences in pre-spawning Chinook salmon performance across scales of biological organization at ecologically relevant temperatures. Given the rapid warming of rivers, we show that it is critical to consider the intra-specific variation in thermal physiology to assist in the conservation and management of Pacific salmon.