Whole cell current-clamp recordings were made from 85 sympathetic preganglionic neurones (SPN) of the neonatal spinal cord in vitro. Superfusion of up to 500 microM acetylcholine (2-30 sec) gave weak responses. Carbachol (CChol; 5-50 microM) superfused for 1-20 sec, hyperpolarized SPN. This response was associated with a mean reduction in input resistance of 22%. Ion substitution studies suggested that potassium is the likely carrier of at least part of the current underlying the CChol-induced hyperpolarization. Some SPN display spontaneous rhythmic oscillations in their membrane potentials which may be due to action potential discharge in electrically-coupled neurones. CChol also acts to inhibit these oscillations concomitant with the hyperpolarization. Responses to CChol were unaffected by addition of 500 nM TTX to the bathing medium suggesting that CChol acts directly upon SPN. Carbachol-induced hyperpolarizing responses were totally abolished by the non-specific muscarinic receptor antagonist atropine (20-50 microM). Pirenzepine, at concentrations over 5 microM reversibly reduced the responses to CChol. Gallamine, an M2 receptor antagonist applied at 25 microM also reversibly abolished the CChol responses. These results suggest that CChol-mediated hyperpolarizations may be due to M2 receptor activation.