1. 5-Hydroxytryptamine (5-HT) was applied by microiontophoresis in the vicinity of identified sympathetic preganglionic neurones in the upper thoracic spinal cord of the rat, in vivo. 2. Sympathetic preganglionic neurones responded in one of three ways to 5-HT: by (a) excitation (76%), (b) inhibition (4%) or (c) in a biphasic manner (5%). 3. The excitatory responses evoked by 5-HT were mimicked by 5-carboxamidotryptamine (5-CT) and alpha-methyl-5-hydroxytryptamine (alpha-Me-5-HT). The inhibitory and biphasic responses evoked by 5-HT were mimicked by 2-methyl-5-hydroxytryptamine (2-Me-5-HT). The observed responses evoked by 5-HT and selective agonists may be different on the same cell. In several instances a single neurone excited by one agonist was inhibited by another agonist. 4. The 5-HT2-receptor antagonists, ketanserin and LY 53857, failed to abolish selectively the excitatory responses evoked by 5-HT and alpha-Me-5-HT, when applied by microiontophoresis. The antagonists non-selectively reduced the excitatory responses evoked by 5-HT, 5-CT, alpha-Me-5-HT, D,L-homocysteic acid (DLH) and noradrenaline (NA). A reduction in synaptically evoked activity was also observed. 5. The 5-HT3-receptor antagonist, ICS 205-930, failed to abolish the inhibitory responses evoked by 5-HT. 6. It was concluded that the excitatory responses evoked by 5-HT are mediated by a receptor that is neither 5-HT2 or 5-HT3, but shows similarities to the 5-HT1-like receptor profile. The inhibitory actions of 5-HT are mimicked by 2-Me-5-HT, but the receptor is not 5-HT3, or 5-HT1-like or 5-HT2.