Specific antibodies raised against a fusion protein containing the amino acid sequence of the putative second intracellular loop of the cloned 5-HT3-A receptor subunit were used for the immunohistochemical visualization of 5-HT3 receptors in the rat spinal cord. A dense 5-HT3-like immunoreactivity was found in the superficial layers of the dorsal horn, which closely matched the labelling of 5-HT3 binding sites by [125I]iodo-zacopride. This immunostaining was markedly decreased following unilateral rhizotomy, consistently with a preferential location of 5-HT3 receptors on terminals of primary afferent fibres, and with the presence of 5-HT3 mRNA in dorsal root ganglia. However, a significant proportion of 5-HT3 receptors persisted after rhizotomy, and the corresponding mRNA was found in the dorsal horn of the spinal cord. 5-HT3 receptors are therefore also located on intrinsic neurones of the spinal cord.