Previous studies employing 5-hydroxydopamine to identify nigrostriatal dopaminergic axons and their synapses found that labeled axons made few synapses or that asymmetric contacts predominated. In contrast, recent studies using tyrosine hydroxylase or dopamine antibody techniques indicate that presumed dopaminergic axons form small symmetric contacts. We re-examined 5-hydroxydopamine-labeled material from the rat neostriatum using serial three-dimensional reconstruction techniques to characterize the morphology of labeled axons, synapses and postsynaptic targets. This ultrastructural analysis revealed a class of heavily labeled axons that are small (0.06-1.5 microns in diameter) and lack large varicosities. These axons form small (0.011-0.09 microns 2), en passant, symmetric synapses, mainly onto dendritic spines and spiny dendritic shafts and, in some cases, onto aspiny dendritic segments near branch points. The sites of these synapses along the axon appeared unrelated to the locations of axonal enlargements, suggesting that counting varicosities may not be an accurate indication of the extent of dopaminergic innervation in the neostriatum. The characteristics of these 5-hydroxydopamine-labeled elements correspond in all respects to axons and synapses identified as dopaminergic by immunohistochemistry in previous studies. In tissue in which all labeled and unlabeled synapses were classified, approximately 9% of all synapses were identified as dopaminergic by this type of label. Three-dimensional reconstructions provided additional insight concerning the interaction of dopaminergic afferents with postsynaptic striatal targets and their relation to other afferents to these neurons. They reveal that a short, unbranched dopaminergic axonal segment can make multiple synapses onto dendritic spines, shafts and branch points of one or more dendrites. In addition, one dendrite can receive contacts from several labeled axons. Dopamine synapses onto spines are always associated with unlabeled, asymmetric synapses onto the same spine. Synapses of various morphologies with a distinctly different, lighter form of labeling were much rarer, and may represent other aminergic afferents to the neostriatum. The presence of this second form of label in earlier 5-hydroxydopamine studies may have contributed to the long-standing controversy over the appearance of dopaminergic synapses examined by different techniques. Our results help to resolve this controversy and confirm that the nigrostriatal projection makes small symmetric synapses with a variety of striatal targets.