Abstract The development of dopaminergic and noradrenergic neurons in dissociation cultures of mesencephalon and rhombencephalon obtained from 18-day-old rat fetuses was characterized by their capacity to take up and release eatecholamines. In both types of cultures, uptake of [ 3H]dopamine and [ 3H]noradrenaline was obtained which could be inhibited by reserpine. Autoradiographic studies demonstrated an almost exclusive neuronal localization of the labeled catecholamines. The transmitters could be released by depolarization with K + in a Ca 2+-dependent manner during the entire cultivation period. In contrast, catecholamine uptake by cultures of neocortex was minimal, could not be inhibited by reserpine, and the accumulated radioactivity could not be released upon depolarization. These points provide evidence for an active accumulation of the exogenous transmitters and for the presence of stimulus-secretion coupling in a distinct population of neurons of both brain stem cultures. Striking differences between the two brain stem cultures concerned their sensitivity to desmethylimipramine and benztropine as well as the time course of the development of the uptake capacity. Desmethylimipramine inhibited the uptake of both catecholamines in rhombencephalic, but not in mesencephalic cultures. The reverse was true for benztropine. It is concluded that cultures of rhombencephalon contain predominantly noradrenergic, and those of mesencephalon dopaminergic cells. Comparison of the uptake behaviour suggested that noradrenergic neurons mature considerably later than dopaminergic neurons. The results show that dissociation cultures of mid- and hindbrain, inspite of their heterogeneous composition, can serve as valuable models for the study of development and function of dopaminergic and noradrenergic neurons, respectively.