Circadian rhythms are intrinsic timing systems that organisms have evolved to adapt to the Earth's rotation, regulating numerous critical physiological processes ranging from sleep to metabolism. In mammals, the "central clock" of this system is located in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN comprises a vast network of neurons, each functioning as an independent "molecular clock." To ensure the stability of the organism's overall rhythm, these individual "clocks" must achieve a high degree of synchronization. Research has identified the neuropeptide vasoactive intestinal polypeptide (VIP) as a key messenger mediating coupling and synchronization among SCN neurons. However, traditional in vitro static culture methods introduce significant perturbations, such as temperature shocks and serum effects. For a long time, these technical bottlenecks have severely constrained our ability to investigate how VIP signals precisely and dynamically regulate the SCN network.

On January 15, 2026, the research groups of Yanyi Huang and Kui Han from the Biomedical Pioneering Innovation Center (BIOPIC) at Peking University, in collaboration with the group of Erquan Zhang from the National Institute of Biological Sciences (NIBS), published a research paper titled "An Automated Organotypic SCN Culture System Revealing Novel Insights into VIP Regulation of Circadian Rhythm" in the journal Advanced Science. The study reports a self-developed, fully automated, and highly stable brain tissue slice culture system, termed "brain-slice-in-a-chamber" (BaSIC). Utilizing this system, the researchers discovered, for the first time under near-physiological conditions, that the neuropeptide VIP can achieve efficient resetting and synchronization of the mammalian master circadian clock through a novel molecular mechanism: inducing the rapid degradation of the core clock protein PER2. This finding opens new avenues for deeply understanding the molecular mechanisms governing circadian rhythm regulation.

The core of the BaSIC system developed by the research team lies in its precision microfluidic design and unique culture mode, which overcome longstanding bottlenecks in traditional ex vivo culture methods, such as temperature fluctuations, serum shock, and contamination risks. In this system, brain tissue slices are placed on a semi-permeable membrane and maintained at an optimized air-liquid interface, rather than being fully submerged in culture medium. This design more closely mimics the in vivo physiological environment, significantly enhancing tissue viability and physiological activity during long-term culture. BaSIC enables the continuous, stable, and programmable perfusion of culture media and various stimuli (such as neuropeptides). Furthermore, the system integrates high-precision temperature and humidity control modules with high-sensitivity optical monitoring interfaces, allowing for the flexible integration of microscopic imaging systems or photomultiplier tubes. This capability facilitates the long-term, uninterrupted tracking of reporter gene expression within the SCN. The BaSIC system provides a stable, reliable, and powerful technical platform for investigating the fine dynamic processes of neural tissue under near-physiological conditions.

Figure 1. Design and working principle of the BaSIC fully automated brain slice culture system.

Leveraging the BaSIC platform, the research team was able to investigate the impact of periodic VIP signals on SCN rhythms under interference-free conditions. When periodic high-concentration VIP pulses were applied to mouse SCN slices carrying the PER2::LUC reporter gene, the researchers observed that, contrary to predictions from the traditional transcription-translation feedback loop (TTFL) model which suggests an increase in expression, Period 2 (PER2) protein levels underwent a rapid decline following each VIP stimulus. Through optical imaging tracking of individual neuronal rhythms, the team discovered that this rapid decline represents a collective synchronization behavior across the entire SCN neuronal network. Regardless of the initial phase of an individual neuron's rhythm (whether at a peak, rising phase, or falling phase), the high-concentration VIP signal synchronously triggered all neurons into a "silent" state characterized by extremely low PER2 protein levels. This process achieves dual synchronization of both phase and amplitude: the "clock hands" (phase) of all neurons are instantaneously calibrated to a common trough starting point, while their previously diverse protein levels (amplitude) converge to a uniform low baseline. This "resetting" effect ensures that the entire SCN neural network can generate a more stable overall rhythmic output in response to strong external stimuli, such as intense light exposure.

Figure 2. Periodic VIP pulse stimulation induces a rapid and synchronized reduction in PER2 protein levels, with the synchronization index approaching 1 at the trough.

To validate this newly discovered biological function, the research team further utilized the auxin-induced degron (AID) system to achieve artificial rapid degradation of PER2 protein within cells. The results demonstrated that this rapid degradation could induce a strong phase reset of the circadian rhythm, an effect highly consistent with the phenomena observed upon VIP stimulation of the SCN. This finding provides compelling evidence that VIP-induced rapid degradation of PER2 is the core molecular mechanism enabling fast and efficient synchronization within the SCN network.

To theoretically elucidate and verify this new mechanism, the team constructed a mathematical model centered on PER2 protein. This model integrated PER2 transcription, translation, phosphorylation modifications, and conventional degradation pathways, while innovatively introducing a "rapid degradation" pathway triggered by VIP signaling. The simulation results perfectly recapitulated the key phenomena observed in the experiments: VIP pulses could instantaneously trigger a sharp decline in PER2 protein levels, ultimately leading to the resetting of the circadian clock phase in synchrony with the VIP stimulus.

Figure 3. Mathematical modeling reveals that VIP-induced rapid degradation of PER2 is the key to phase resetting.

In summary, this study achieves breakthroughs on two fronts. Technologically, the successful development of BaSIC, an automated in vitro brain tissue research platform, provides a novel tool for neuroscience, particularly for investigating the dynamic processes of brain tissue over extended periods. Scientifically, by integrating experimental observations with theoretical modeling, the study elucidates for the first time a new molecular mechanism whereby VIP signaling forcibly "resets" the circadian clock by inducing the rapid degradation of PER2 protein. This discovery not only significantly deepens our understanding of the mammalian circadian regulatory network but also offers new potential targets and a crucial theoretical foundation for developing future intervention strategies against circadian rhythm disorders.

Professor Yanyi Huang from Peking University, Professor Erquan Zhang from the National Institute of Biological Sciences (NIBS), and Dr. Kui Han from Peking University (currently an Associate Researcher at the Changping Laboratory) serve as the co-corresponding authors of this paper. Dr. Kui Han, Dr. Meimei Liao from Chongqing Medical University, and Jingpeng Zhang, a doctoral student at Peking University, are the co-first authors. Ruoyu Zhong from Peking University, Dr. Long Mei from NIBS, and Dr. Dapeng Ju from Chongqing Medical University made significant contributions to platform construction, cell line generation, and data analysis. This research was supported by National Natural Science Foundation of China, Beijing National Laboratory for Molecular Sciences, National Key R&D Program of China, and STI2030-Major Project.

Link: https://advanced.onlinelibrary.wiley.com/doi/10.1002/advs.202511069