The effects of rat serum extracts on the uptake of [3H]-serotonin and the displacement of [3H]-imipramine binding in rat forebrain synaptosomes and human platelets was studied. Deproteinated rat serum markedly inhibited synaptosomal [3H]-serotonin uptake in a dose-dependent and reversible manner. The crude extract was fractionated by C18-reverse phase HPLC. Three major peaks of inhibitory activity were found. One of the peaks was identified as serotonin and was significantly reduced after chronic reserpinization. The second major peak inhibited both [3H]-serotonin uptake and [3H]-imipramine binding in synaptosomes and platelets. This fraction had a minimal effect on the uptake of [3H]-norepinephrine, [3H]-dopamine or [3H]-GABA and was less effective in inhibiting [3H]-desipramine binding than [3H]-imipramine binding.