The cerebellum receives indolaminergic fibers influencing Purkinje cell discharges. Data from our laboratories have demonstrated an endogenous release of serotonin (5-HT) and a Na(+)-dependent uptake and Ca(2+)-dependent release of [3H]5-HT from slices, homogenates and synaptosomal fractions of the rat cerebellar molecular layer. While the neurotransmitter produced by climbing fibers has been sought for in several studies and some of the classical transmitters have been ruled out, as yet this neurotransmitter is unknown. The aim of this work was to measure the 5-HT uptake and release from rat cerebellar slices, 6 h and 15 days after intraperitoneal injection of 3-acetylpyridine (3-AP) (75 mg/kg), harmaline (15 mg/kg) and nicotinamide (300 mg/kg). A histological study of medulla and cerebellar cortex in these animals showed destruction of neurons in the inferior olivary nuclei and changes in the granulation of the cortical molecular layer in the cerebellum. A significant reduction of the 5-HT content (100%), 5-HT uptake (60%) and its Vmax (60%) was seen on the 5th day, in cerebellar preparations obtained from rats injected with 3-AP. The Ca(2+)-dependent release of 5-HT from these preparations was found to be similar to the basal values, in spite of depolarizing stimuli with 53 mM KCl or veratrine (60 micrograms/ml). The results suggest that 5-HT could play an important role as neurotransmitter produced by some climbing fibers.