Intracellular recordings were made from rat dorsal raphe neurons in vitro. Baclofen (30 microM) and 5-carboxamidotryptamine (5-CT, 300 nM to 1 microM) hyperpolarized these neurons by 10 and 13 mV, respectively. Depolarizing synaptic potentials (DSPs) were evoked by single shocks: baclofen reduced the amplitude of the DSP by 81%, but 5-CT reduced it by only 23%. The somatic response to iontophoretically applied glutamate pulses was reduced by 12% by baclofen, and 23% by 5-CT. In slices from rats pretreated with intracerebroventricular pertussis toxin (PTX), the ability of baclofen to reduce the DSP was almost unchanged, although the hyperpolarizing action of baclofen, and both actions of 5-CT were virtually eliminated. We conclude that it is possible to distinguish the pre- and postsynaptic actions of baclofen with PTX, and that the actions of 5-CT are both blocked.