The Choline acetyltransferase (ChAT) activity of rat cerebral cortex, cerebellum, hippocampus, hypothalamus midbrain, pons, medulla oblongata and caudate nucleus was determined in control, phenobarbital-treated and picrotoxin-treated animals. 2. Phenobarbital increased ChAT activity in the cerebral cortex an caudate nucleus only. 3. Picrotoxin increased ChAT activity in the cerebellum and caudate nucleus. 4. These findings further clarify the effects of CNS depressants and stimulants on rodent brain cholinergic system.