1. An in vitro brain stem slice from adult guinea pigs was used to study the effects of membrane hyperpolarization in two classes of bulbospinal neurons, called types I and II, from the ventral parts of the nucleus tractus solitarius (vNTS). These bulbospinal neurons project to the phrenic motor nucleus and make up the dorsal respiratory group, a sensorimotor integrating area for rhythmic breathing movements. 2. Negative current injections (1 s long) were used in the discontinuous current-clamp mode to study the input resistance (Rin) in both classes of bulbospinal vNTS neurons. The mean Rin for type I neurons was 88.7 +/- 13.8 (SD) M omega (n = 19) and for type II neurons was 92.6 +/- 14.0 M omega (n = 16). Both classes of neurons displayed a depolarizing sag and inward rectification during negative current injections to membrane-potential levels less than or equal to -70 mV. The magnitude of the depolarizing sag became larger as the size of the negative current step was increased. On release from hyperpolarization, both cell types also exhibited a large anode break hyperpolarization (ABH). 3. The ABH was abolished in the presence of 5 mM 4-amino-pyridine (4-AP), whereas the depolarizing sag and inward rectification were not affected. In the place of the ABH, a small postinhibitory rebound (PIR) depolarization was observed on release from hyperpolarization. The magnitude of PIR was dependent on the size of the depolarizing sag. In the presence of both 5 mM 4-AP and 5 mM Cs+, the depolarizing sag and PIR were completely blocked, whereas Rin was increased. 4. The ionic currents underlying the ABH and depolarizing sag were directly observed by the use of the discontinuous single-electrode voltage-clamp technique. The ABH was caused by activation of an A-current (IKA). The depolarizing sag was associated with a hyperpolarization-activated inward current (IH), which was activated at membrane-potential levels less than or equal to -70 mV. The peak amplitude of IH in type I neurons was -335 +/- 16 pA (n = 13) and in type II cells was -327 +/- 14 pA (n = 11). 5. IH currents did not display inactivation during the hyperpolarizing voltage step. The IH current became larger when [K+]o was increased from 4 mM (control) to 12 mM and was blocked in the presence of 5 mM Cs+. The estimated reversal potential for the IH current was -41.5 +/- 4.8 mV (n = 8).(ABSTRACT TRUNCATED AT 400 WORDS)