Using 7.4 kb of 5' flanking DNA from the Drosophila cholinergic gene locus to drive Gal4 expression we can visualize essentially all cholinergic neurons and neuropiles after genetic recombination with a UAS-GFP (S65T) reporter gene. In contrast to previous methods somata and neuropiles can be observed in the same samples. Fluorescence intensity is strong enough to allow observations in live animals at all developmental stages. Three-dimensional reconstructions made from confocal sections of whole-mount preparations reveal the extensive cholinergic connections among various regions of the nervous system.