A crude preparation of neurotransmitter storage vesicles was obtained by differential centrifugation and the ability to take up 3H-dopamine was evaluated in vitro. The uptake was highly dependent on temperature, had an absolute requirement for ATP and Mg2+ and was inhibited totally by reserpine. The uptake displayed saturation kinetics, with a Km of 0.26 μM at 20°. 3H-dopamine uptake was inhibited competitively by norepinephrine, with a Ki of 0.69 μM. Vesicles derived from a primarily dopaminergic region (corpus striatum) exhibited the same ratio of uptakes of 3H-dopamine/3H-norepinephrine as did those from a primarily noradrenergic region (cerebral cortex). These results indicate that viable rat brain storage vesicles can be readily prepared and used for evaluation of pharmacologic effects on 3H-dopamine uptake, and that dopaminergic and noradrenergic storage vesicles exhibit identical uptake properties. © 1978.