Mapping of circadian clock-related neuron connections in Drosophila

The circadian clock, an internal timekeeping system, enables organisms to respond to rhythmic environmental changes occurring over 24 hours. The master circadian clock located in the brain regulates several peripheral clocks in different tissues through endocrine and systemic signaling. These neurons, along with neurotransmitters, neuropeptides, and gap junctions, form a neuronal network. The circadian clock neural network is highly conserved across species, making the fruit fly Drosophila melanogaster an ideal model for circadian clock studies.

A collaborative study led by Professor Taishi Yoshii from Okayama University, Nils Reinhard and Charlotte Helfrich-Förster from the University of Würzburg, and Meet Zandawala from the University of Nevada Reno has made a breakthrough in understanding the regulation of rhythmic physiology through the circadian clock. The team has constructed a comprehensive map of the synaptic connectome of clock neurons in Drosophila. The study was published online on December 05, 2024, in the Journal Nature Communications. Prof. Yoshii said, “The connectome developed in this study revealed the importance of clock neurons in regulating the feeding and mating behaviors of Drosophila.” Using the FlyWire connectome, the researchers identified 242 clock neurons based on morphology, known connectivity, and location of their soma. Prof. Yoshii said, “The number of clock neurons identified in this study is higher than the previously reported 150 neurons, indicating the presence of novel neurons.” The study also identified new neuron subtypes, including dorsal neurons (DN₃ and DN_1p), and connections among different subgroups across both brain hemispheres. Particularly, DN_1pA was identified as an important center linking the clock networks across the two brain hemispheres as it formed both ipsilateral and contralateral connections. This extensive contralateral synaptic connectivity indicates that the neurons on the opposite sides of the brain are connected.

The study revealed that light input pathways influence clock neurons. Photoreceptor cells of the compound eyes, Hofbauer-Buchner eyelets, and ocelli provided indirect inputs to the clock neurons, indicating a new layer of complexity in how light stimulus plays an important role in the functioning of the circadian neurons.

Furthermore, the study suggests that the output of the clock network neurons was directed toward the intrinsic brain neurons, especially central brain neurons. The clock network utilized descending neurons, such as Allostatin-C and SIFamine (SIFa) peptidergic neurons, which are involved in regulating feeding, mating, and sleep behaviors.

Neurotransmitter analysis showed that most of the lateral clock neurons expressed the excitatory neurotransmitter acetylcholine, whereas most of the dorsal clock neurons expressed glutamate. In remarks on this, Prof. Yoshii explained, “The inhibitory neurotransmitter GABA was absent in clock neurons. Glutamatergic (small-central projecting DN₃) and cholinergic clock neurons were involved in synaptic outputs to neurons that regulate hunger, thirst, and mating behaviors.”

Additionally, the study identified 12 neuropeptides that were upregulated in the clock network. Among these neuropeptides, DH44 and Proctolin were the novel neuropeptides identified in this study. This suggests that clock neurons communicate through peptidergic paracrine signaling.

These findings significantly improved our understanding of the connectivity and functioning of clock neurons and have potential implications for the development of therapeutic strategies for circadian disorders, such as sleep and mood disorders in humans.

About Okayama University, Japan

As one of the leading universities in Japan, Okayama University aims to create and establish a new paradigm for the sustainable development of the world. Okayama University offers a wide range of academic fields, which become the basis of integrated graduate schools. This not only allows us to conduct the most advanced and up-to-date research, but also provides an enriching educational experience.

Website: https://www.okayama-u.ac.jp/index_e.html

About Professor Taishi Yoshii from Okayama University, Japan

Dr. Taishi Yoshii has been a Professor at the Graduate School of Environmental, Life, Natural Science and Technology, Okayama University, Japan, since April 2022. He completed his PhD from Okayama University in 2006 and his Master’s degree from Yamaguchi University, Yamaguchi, Japan, in 2003. He was awarded the Excellent Poster Award at the 19^th Academic Conference of the Japanese Chronobiology Society. He has published his research in various peer-reviewed journals, such as the Journal of Biological Rhythms, Cell Reports, and Biochemical and Biophysical Research Communications. His research interests are chronobiology, circadian rhythms, and Drosophila melanogaster.