GFP labeling of neurosecretory cells with the GAL4/UAS system in the silkmoth brain enables selective intracellular staining of neurons

The microbrain of the silkmoth, Bombyx mori, is a model system for analyzing the neural mechanisms underlying stimulus-driven behavior, and numerous studies using physiological and morphological methods have accumulated. However, one of the limitations of this system is a lack of methodology for labeling specific subsets of neurons. Targeted gene expression with the GAL4/UAS system, which was recently developed, may overcome this disadvantage. To test the GAL4/UAS system in the silkmoth brain, we generated two GAL4 driver lines in which GAL4 expression was under the control of either the bombyxin or prothoracicotropic hormone (PTTH) promoter. Crosses of moths from these lines with a UAS-GFP line showed that green fluorescent protein (GFP) was exclusively expressed in bombyxin or PTTH neurosecretory brain cells. Using these lines, we developed a visually guided method to selectively insert an electrode into and intracellulary stain GFP-expressing cells using fluorescence as a landmark. This work provides a novel method to visualize specific subsets of neurons in the silkmoth brain and to observe detailed structures in a single identified neuron from different individuals.