Abundances of coldwater adapted stream fish populations are declining largely due to anthropogenic influences, including increased temperature. To persist in streams with unsuitable thermal habitat, fish must move to coldwater patches, acclimate, or adapt to water temperatures above thermal optima. Brook trout, a coldwater adapted salmonid, has previously displayed physiological plasticity and the ability for reversible thermal acclimation when reared at higher temperatures. However, because stream temperatures are not static, it is important to explore the rate at which thermal acclimation occurs to evaluate whether prior thermal experience will influence future thermal performance. To determine the temporal scale in loss of thermal acclimation as water temperatures cool, we acclimated brook trout to three thermal regimes: +0 °C (ambient; mimicking the daily average water temperature of a nearby long-term study site), as well as +2 °C and +4 °C above ambient. After 2 years of being reared under those conditions, fish from the warmer treatments were moved to a common, colder temperature (ambient). We then used critical thermal maximum to measure the loss in acclimation response of fish from each treatment over time. We found that regardless of initial acclimation temperature, thermal tolerance of warm acclimated fish decreased rapidly for 1 week, then gradually decreased, and was completely lost within 42 days. This gradual loss of acclimation may be valuable to persistence in warmer streams and will be important to include in models of the impact climate change has on brook trout and other aquatic ectotherms with significant thermal plasticity.