Electrophysiological studies using rats anesthetized with chloral hydrate were performed to determine whether or not serotonin originating in the dorsal raphe nucleus (DR) acts as an inhibitory transmitter or neuromodulator on neurons of the dorsal lateral geniculate nucleus (LGN) and neurons located in the thalamic reticular nucleus (TRN) immediately rostral to the dorsal LGN. In the LGN, conditioning stimuli applied to the DR preceding test stimulus to the optic tract and visual cortex inhibited orthodromic and antidromic spikes in about one-third of the relay neurons and in more than half of the intrageniculate interneurons. Conditioning stimulation of the DR also produced an inhibition of the spikes elicited by stimulation of the optic tract and visual cortex of at least three-quarters of the TRN neurons. Iontophoretic application of serotonin (25 nA) inhibited the orthodromic spikes of the LGN relay neuron and TRN neuron. A close correlation was observed between the effects of DR conditioning stimulation and iontophoretic serotonin in the same neurons. The inhibition with DR conditioning stimulation and iontophoretically applied serotonin was antagonized during iontophoretic application of methysergide (15-40 nA), a serotonin antagonist. These results strongly suggest that serotonin derived from the DR acts on the LGN and TRN neurons as an inhibitory transmitter or neuromodulator to inhibit transmission in these nuclei.