Marked changes were revealed in the activity of the key enzyme in serotonin biosynthesis, tryptophan hydroxylase (TPH), during entry into hibernation, hibernation, and arousal in ground squirrels (Citellus erythrogenys). An increase in TPH activity was found in the midbrain, hippocampus, and striatum during the prehibernation period in euthermic ground squirrels. A further increase in TPH activity was observed during the entry into hibernation. Significant elevation was found not only in potential TPH activity measured at the incubation temperature of 37 degrees C but also at incubation temperature of 7 degrees C, approximating the body temperature in hibernation. Vmax in the midbrain of hibernating animals was about 50% higher than in active ones without significant changes in Km. Thus, brain TPH maintains functionality during torpidity and is activated before the entry into hibernation. The results support the idea that brain serotonin is crucially involved in the transition to and the maintenance of the hibernation state.