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Complex behaviors arise from neural circuits that assemble from diverse cell types. Sleep is a conserved behavior essential for survival, yet little is known about how the nervous system generates neuron types of a sleep-wake circuit. Here, we focus on the specification of Drosophila 23E10-labeled dorsal fan-shaped body (dFB) long-field tangential input neurons that project to the dorsal layers of the fan-shaped body neuropil in the central complex. We use lineage analysis and genetic birth dating to identify two bilateral type II neural stem cells (NSCs) that generate 23E10 dFB neurons. We show that adult 23E10 dFB neurons express ecdysone-induced protein 93 (E93) and that loss of ecdysone signaling or E93 in type II NSCs results in their misspecification. Finally, we show that E93 knockdown in type II NSCs impairs adult sleep behavior. Our results provide insight into how extrinsic hormonal signaling acts on NSCs to generate the neuronal diversity required for adult sleep behavior. These findings suggest that some adult sleep disorders might derive from defects in stem cell-specific temporal neurodevelopmental programs.
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This content will become publicly available on April 1, 2026
Lineage Analysis and Birthdating of Drosophila Central Complex Lineages
From insects to humans, the nervous system generates complex behaviors mediated by distinct neural circuits that are composed of diverse cell types. During development, the spatiotemporal gene expression of the neural progenitors expands the diversity of neuronal and glial subtypes. Various neural stem cell–intrinsic and –extrinsic gene programs have been identified that are thought to play a major role in generating diverse neuronal and glial cell types.Drosophilahas served as an excellent model system for discovering the fundamental principles of nervous system development and function. The sophisticated genetic tools allow us to link the origin and birth timing (the time when a particular neuron is born during development) of neuron types to unique neural stem cells (NSCs) and to a developmental time. InDrosophila, a special class of NSCs called Type II NSCs has adopted a more advanced division mode to generate lineages for the higher-order brain center, the central complex, which is an evolutionarily conserved brain region found in all insects. Type II NSCs, similar to the human outer radial glia, generate intermediate neural progenitors (INPs), which divide many times to produce about eight to 10 progeny. Both Type II NSCs and INPs express distinct transcription factors and RNA-binding proteins that have been proposed to regulate the specification of cell types populating the adult central complex. Here, we describe the recently invented lineage filtering system, called cell class–lineage intersection (CLIn), which enables the tracking and birthdating of the Type II NSC lineages. Using CLIn, one can easily generate clones of different Type II NSCs and identify not only the origins of neurons of interest but also their birth time.
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- Award ID(s):
- 2047020
- PAR ID:
- 10625213
- Publisher / Repository:
- CSHL Press
- Date Published:
- Journal Name:
- Cold Spring Harbor Protocols
- Volume:
- 2025
- Issue:
- 4
- ISSN:
- 1940-3402
- Page Range / eLocation ID:
- pdb.prot108442
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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