Sleep Aid Disrupts Norepinephrine-Mediated Waste Clearance in Mice

A good night’s sleep does more than just help you feel rested, it might literally clear your mind, according to the results of newly reported research by scientists at the University of Rochester and at the University of Copenhagen. Their study in mice showed how deep sleep may wash away waste buildup in the brain during waking hours, an essential process for maintaining brain health.

The results for the first time describe the tightly synchronized oscillations in the neurotransmitter norepinephrine (NE), cerebral blood, and cerebrospinal fluid (CSF) that combine during non-rapid eye movement (NREM) sleep in mice. These oscillations power the glymphatic system, a brain-wide network that is responsible for removing protein waste, including amyloid and tau that are associated with neurodegenerative diseases.

The study also presents new insights into how sleep aids may disrupt the “brainwashing” system, potentially affecting cognitive function over the longer term, and holding a warning for people who use the commonly prescribed sleep aid zolpidem. The results indicated that in mice the drug suppressed the glymphatic system, potentially setting the stage for neurological disorders such as Alzheimer’s disease, which are the result of the toxic accumulation of proteins in the brain.

“As the brain transitions from wakefulness to sleep, processing of external information diminishes while processes such as glymphatic removal of waste products are activated,” said Maiken Nedergaard, MD, DMSc, co-director of the University of Rochester Center for Translational Neuromedicine. “The motivation for this research was to better understand what drives glymphatic flow during sleep, and the insights from this study have broad implications for understanding the components of restorative sleep.”

Nedergaard is lead author of the researchers’ published study in Cell, titled “Norepinephrine-mediated slow vasomotion drives glymphatic clearance during sleep.”

Scientists have known that the brain has a built-in waste removal system, called the glymphatic system, which circulates fluid in the brain and spinal cord to clear out waste. This process helps remove toxic proteins that form sticky plaques linked to neurological disorders. Glymphatic fluid transport is also enhanced during sleep, the researchers wrote. “As the brain transitions from wakefulness to sleep, processing of external information diminishes while restorative processes, such as glymphatic removal of waste products, are activated.”

In recent years the link between sleep and the glymphatic system has received substantial attention, as poor sleep often precedes the onset of neurodegenerative diseases and is a predictor of early dementia, the team continued. “Yet, the precise mechanisms by which sleep and wakefulness influence glymphatic flow remain unclear.”

For their newly reported study in mice, the researchers employed an optic technique called flow fiber photometry, combined with electroencephalogram and electromyography monitors. Unlike previous research techniques, which immobilized the mice and used anesthesia to induce sleep, the new approach enabled researchers to record brain activity during long, uninterrupted periods of wakefulness and sleep, while allowing mice to move freely during recordings. The team explained, “We here developed a method, ‘flow fiber photometry,’ that enabled recordings of blood and CSF dynamics during long, uninterrupted periods of wakefulness, NREM, and REM sleep by avoiding the need for head fixation and allowing mice to move freely in their home cage during recordings.”

Their findings highlighted the critical role of norepinephrine, a neurotransmitter associated with arousal, attention, and the body’s response to stress. During deep sleep, the brainstem releases tiny waves of norepinephrine about once every 50 seconds. Norepinephrine triggers blood vessels to contract, generating slow pulsations that create a rhythmic flow in the surrounding fluid to carry waste away.

“It’s like turning on the dishwasher before you go to bed and waking up with a clean brain,” said Nedergaard “We’re essentially asking what drives this process and trying to define restorative sleep based on glymphatic clearance.”

Nedergaard and her team looked more closely at what happens in mice when the brain sleeps. Specifically, they focused on the relationship between norepinephrine and blood flow during deep slumber. They found that norepinephrine waves correlated with variations in brain blood volume, suggesting that norepinephrine triggers a rhythmic pulsation in the blood vessels.

The team observed that slow synchronized waves of norepinephrine, cerebral blood volume, and CSF flow characterized NREM sleep. The norepinephrine triggered “micro-arousals,” causing vasomotion, the rhythmic constriction of blood vessels independent of the heartbeat. This oscillation, in turn, generates the pumping action necessary to move CSF in the glymphatic system during sleep.

“These findings, combined with what we know about the glymphatic system, paint the whole picture of the dynamics inside the brain, and these slow waves, micro-arousals, and the norepinephrine were the missing link,” said Natalie Hauglund, PhD, first author of the study and currently a postdoctoral fellow at the University of Oxford. “You can view norepinephrine as this conductor of an orchestra. There’s a harmony in the constriction and dilation of the arteries, which then drives the cerebrospinal fluid through the brain to remove the waste products.”

In their paper, the authors pointed out, “The results presented here reinforce previous studies that identified sleep as a key driver of glymphatic flow in the rodent and human brain.” Added Nedergaard, “Now we know norepinephrine is driving the cleaning of the brain, we may figure out how to get people a long and restorative sleep.”

The study also explored whether sleep aids replicate the natural oscillations necessary for glymphatic function. The team focused on zolpidem, a sedative that is commonly prescribed to treat insomnia.

They found that while zolpidem effectively induced sleep in the mice, it also suppressed norepinephrine oscillations, disrupting the glymphatic system and impeding the brain’s waste-clearing processes, a finding that may raise concerns about its long-term use. Norepinephrine waves during deep sleep were 50% lower in zolpidem-treated mice than in naturally sleeping mice. Although the zolpidem-treated mice fell asleep faster, fluid transport into the brain dropped more than 30%. “Our analysis showed that zolpidem impaired NE oscillations and reduced glymphatic CSF tracer inflow,” the investigators wrote. The findings suggest that the sleeping aid may disrupt the norepinephrine-driven waste clearance during sleep.

“More and more people are using sleep medication, and it’s really important to know if that’s healthy sleep,” said Hauglund. “If people aren’t getting the full benefits of sleep, they should be aware of that so they can make informed decisions.”

The team said that while further research will be needed, their findings will likely apply to humans, who also have a glymphatic system. Researchers have observed similar norepinephrine waves, blood flow patterns, and brain fluid flux in humans. The collective findings may offer insights into how poor sleep may contribute to neurological disorders such as Alzheimer’s disease.

Scientists also now have a new tool and potential target to improve sleep. “The research provides a mechanistic link between norepinephrine dynamics, vascular activity, and glymphatic clearance, advancing understanding of sleep’s restorative functions,” said Nedergaard. “It also calls attention to the potentially detrimental effects of certain pharmacological sleep aids on brain health, highlighting the necessity of preserving natural sleep architecture for optimal brain function.”

In their paper, the scientists stated that their results “… have broad implications for understanding the components of restorative sleep, the functions of slow vasomotion, and the connection between vascular dysfunction and glymphatic failure, which predispose individuals to neurodegenerative diseases.”

The authors further wrote, “Clinically, zolpidem has been associated with increased risk of reversible dementia and linked to neuropsychiatric adverse events … our study indicates that while zolpidem reduces the latency to sleep, it interferes with normal sleep architecture and suppresses glymphatic flow in mice. This suggests that sleep aids like zolpidem are unlikely to improve glymphatic function, aligning with the general consensus that these drugs carry considerable adverse effects and are associated with increased mortality risk.”