Neurotransmitter and Receptor Mapping in Drosophila Circadian Clock Neurons via T2A-GAL4 Screening

The circadian neuronal network in the brain comprises central pacemaker neurons and associated input and output pathways. These components work together to generate coherent rhythmicity, synchronize with environmental time cues, and convey circadian information to downstream neurons that regulate behaviors such as the sleep/wake cycle. To mediate these functions, neurotransmitters and neuromodulators play essential roles in transmitting and modulating signals between neurons. In Drosophila melanogaster, approximately 240 brain neurons function as clock neurons. Previous studies have identified several neurotransmitters and neuromodulators, including the Pigment-dispersing factor (PDF) neuropeptide, along with their corresponding receptors in clock neurons. However, our understanding of the neurotransmitters and receptors involved in the circadian system remains incomplete. In this study, we conducted a T2A-GAL4-based screening for neurotransmitter and receptor genes expressed in clock neurons. We identified 2 neurotransmitter-related genes and 22 receptor genes. Notably, while previous studies had reported the expression of 6 neuropeptide receptor genes in large ventrolateral neurons (l-LN_v), we also found that 14 receptor genes-including those for dopamine, serotonin, and γ-aminobutyric acid-are expressed in l-LN_v neurons. These findings suggest that l-LN_v neurons serve as key integrative hubs within the circadian network, receiving diverse external signals.