1. Intracellular recordings were made from antidromically identified sympathetic preganglionic neurones (SPNs) in transverse sections of thoraco-lumbar spinal cord from neonatal (12-22 day) rats. 2. Two types of hyperpolarizing (inhibitory) postsynaptic potentials (IPSPs) were recorded in the SPNs. The first type, which we have termed unitary IPSPs, were small, discrete IPSPs that occurred spontaneously and also following chemical or electrical stimulation applied to the spinal cord slices. The second type IPSP was a hyperpolarizing response evoked by either dorsal or ventral root stimulation. 3. Spontaneously occurring unitary IPSPs had an amplitude of 1 to 5 mV, and reversal potential of -60 to -75 mV; they were reversibly abolished by low Ca2+, tetrodotoxin (TTX) or strychnine but not by bicuculline and picrotoxin. 4. Pressure application of N-methyl-D-aspartate (NMDA), an excitatory amino SPNs; these were abolished by either strychnine or by the NMDA receptor antagonist D-2-amino-5-phosphonovalerate. Furthermore, electrical stimulation of dorsal rootlets elicited in several SPNs the discharge of strychnine-sensitive unitary IPSPs. 5. Electrical stimulation applied to dorsal or ventral rootlets elicited in nineteen and eight SPNs, respectively, an IPSP of larger amplitude (5 to 15 mV). The IPSP exhibited a reversal potential of -60 to 75 mV; it was changed to a depolarizing response in a low [Cl-]o solution, but was not significantly affected in a low [K+]o. Strychnine but not bicuculline or picrotoxin reversibly blocked the IPSPs in nearly all the SPNs. Additionally, hexamethonium and d-tubocurarine antagonized the IPSPs evoked by ventral but not by dorsal root stimulations. 6. Our results suggest that unitary and evoked IPSPs recorded in SPNs are due primarily to an increase of Cl- conductance by glycine or a glycine-like substance, released from interneurones, that can be activated by NMDA. Furthermore, IPSPs evoked by ventral root stimulation appear to represent a disynaptic event whereby nicotinic activation of a glycine-releasing interneurone results in a release of the inhibitory transmitter; this is then analogous to the Renshaw cell circuitry of the spinal motoneurones.