40 Hz Light Flickering Promotes Sleep through Cortical Adenosine Signaling

Flickering light stimulation has emerged as a promising non-invasive neuromodulation strategy to alleviate neuropsychiatric disorders. However, the lack of a neurochemical underpinning has hampered its therapeutic development. Here, we demonstrate that light flickering triggered an immediate and sustained increase (up to 3 hours after flickering) in extracellular adenosine levels in the primary visual cortex and other brain regions, as a function of light frequency, intensity, and wavelength, with maximal effects observed at 40 Hz frequency. We discovered cortical (glutamatergic and GABAergic) neurons, rather than astrocytes, as the cellular source, and intracellular adenosine generation from calcium influx-triggered, AMPK- associated energy metabolism pathways (but not SAM-transmethylation or salvage purine pathways) and adenosine efflux mediated by equilibrative nucleoside transporter-2 (ENT2) as the molecular pathway responsible for extracellular adenosine generation. Importantly, 40 Hz light flickering for 30 min enhanced sleep in mice in a frequency-dependent manner. This somnogenic effect was absent in mice lacking ENT2 but replicated by administering adenosine to the visual cortex. Brief 40 Hz light flickering also promoted sleep in children with insomnia by decreasing sleep onset latency, increasing total sleep time, and reducing waking after sleep onset. Collectively, our findings establish adenosine signaling via ENT2 as the neurochemical basis for 40 Hz flickering-induced sleep and unravel a novel and non-invasive treatment for insomnia, a condition that affects 20% of the world population.