1. The thermal adaptation hypothesis proposes that because thermoregulation involves a high metabolic cost, thermal limits of organisms must be locally adapted to temperatures experienced in their environments. There is evidence that tolerance to high temperatures decreases in insects inhabiting colder habitats and microclimates. However, it is not clear if thermal limits of ectotherms with contrasting temporal regimes, such as diurnal and nocturnal insects, are also adapted to temperatures associated with their circadian activities. 2. This study explores differences in heat tolerance among diurnal and nocturnal ant species in four ecosystems in Mexico: tropical montane, tropical rainforest, subtropical dry forests, and high-elevation semi-desert. 3. The critical thermal maximum (CTmax), i.e. the temperature at which ants lost motor control, was estimated for diurnal and nocturnal species. CTmax for 19 diurnal and 12 nocturnal ant species distributed among 45 populations was also estimated. 4. Semi-desert and subtropical dry forest ants displayed higher tolerances to high temperatures than did ants in tropical rainforest. The lowest tolerance to high temperatures was recorded in tropical montane forest ants. In general, among all habitats, the CTmax of nocturnal ants was lower than that of diurnal ants. 5. An increase in nocturnal temperatures, combined with lower tolerance to high temperatures, may represent a substantial challenge for nocturnal ectotherms in a warming world. © 2017 The Royal Entomological Society