1. Intracellular recordings were made from neurones of the nucleus prepositus hypoglossi (PH) in slices of guinea-pig brain. Focal stimulation evoked an inhibitory postsynaptic potential (IPSP) that was typically 10-25 mV in amplitude and 1 s in duration. The IPSP reversal potential showed a Nernstian dependence on the external potassium concentration ([K+]o). 2. Spiperone blocked the IPSP with an IC50 of 40 nM, while ketanserin and (-)sulpiride had no effect. Cocaine (1 microM) prolonged the IPSP half-duration by 157%, and increased the amplitude by 28%. 3. 5-Hydroxytryptamine (5-HT, serotonin) hyperpolarized PH cells with an EC50 of 8.5 microM in control, and 135 nM in cocaine (10 microM). 8-Hydroxy-2-(di-n-propylamino)-tetralin (8-OH-DPAT) also hyperpolarized PH cells with an EC50 of 16 nM, although the maximal effect was only 81% of the maximum 5-HT hyperpolarization. Spiperone produced a parallel, right shift of the 5-HT concentration-response curve; Schild analysis gave a Kd of 10 nM. Application of 5-HT to neurones voltage-clamped near their resting potential (about -55 mV) caused an outward current and an increase in membrane conductance. 4. The amplitude of the IPSP was reversibly decreased by non-hyperpolarizing concentrations of 5-HT and by the 5-HT1 receptor agonists 1-(m-trifluoromethylphenyl)piperazine (TFMPP) and 1-(3-chlorophenyl)piperazine (mCPP). The IC50 values for the latter two compounds were 50 nM and 1.5 microM, respectively; the maximal effect was a 90% inhibition. Neither compound affected the membrane potential nor changed the hyperpolarization induced by 5-HT. Quipizine competitively antagonized TFMPP with an estimated Kd of 165 nM. 5. When trains of stimuli were applied, an inhibition of the IPSP was observed following the first stimulus. At a frequency of 1 Hz, the inhibition was approximately 75%. This frequency-dependent 'run-down' of the IPSP was markedly attenuated by pre-treatment with TFMPP (1 microM). 6. It is concluded that the IPSP in PH cells is caused by 5-HT acting on 5-HT1A receptors to activate a potassium conductance. The release of 5-HT can be inhibited by activation of a presynaptic 5-HT1D receptor. This presynaptic receptor appears to be at least partly responsible for the run-down phenomenon, and may be involved in the physiological regulation of 5-HT synaptic transmission.