Circadian control of brain glymphatic and lymphatic fluid flow.

The glymphatic system is a network of perivascular spaces that promotes movement of cerebrospinal fluid (CSF) into the brain and clearance of metabolic waste. This fluid transport system is supported by the water channel aquaporin-4 (AQP4) localized to vascular endfeet of astrocytes. The glymphatic system is more effective during sleep, but whether sleep timing promotes glymphatic function remains unknown. We here show glymphatic influx and clearance exhibit endogenous, circadian rhythms peaking during the mid-rest phase of mice. Drainage of CSF from the cisterna magna to the lymph nodes exhibits daily variation opposite to glymphatic influx, suggesting distribution of CSF throughout the animal depends on time-of-day. The perivascular polarization of AQP4 is highest during the rest phase and loss of AQP4 eliminates the day-night difference in both glymphatic influx and drainage to the lymph nodes. We conclude that CSF distribution is under circadian control and that AQP4 supports this rhythm.

Biological sex does not predict glymphatic influx in healthy young, middle aged or old mice.

Sexual dimorphism is evident in brain structure, size, and function throughout multiple species. Here, we tested whether cerebrospinal fluid entry into the glymphatic system, a network of perivascular fluid transport that clears metabolic waste from the brain, was altered between male and female mice. We analyze glymphatic influx in 244 young reproductive age (2-4 months) C57BL/6 mice. We found no male/female differences in total influx under anesthesia, or across the anterior/posterior axis of the brain. Circadian-dependent changes in glymphatic influx under ketamine/xylazine anesthesia were not altered by sex. This was not true for diurnal rhythms under pentobarbital and avertin, but both still showed daily oscillations independent of biological sex. Finally, although glymphatic influx decreases with age there was no sex difference in total influx or subregion-dependent tracer distribution in 17 middle aged (9-10 months) and 36 old (22-24 months) mice. Overall, in healthy adult C57BL/6 mice we could not detect male/female differences in glymphatic influx. This finding contrasts the gender differences in common neurodegenerative diseases. We propose that additional sex-dependent co-morbidities, such as chronic stress, protein misfolding, traumatic brain injury or other pathological mechanisms may explain the increased risk for developing proteinopathies rather than pre-existing suppression of glymphatic influx.

Increased glymphatic influx is correlated with high EEG delta power and low heart rate in mice under anesthesia.

The glymphatic system is responsible for brain-wide delivery of nutrients and clearance of waste via influx of cerebrospinal fluid (CSF) alongside perivascular spaces and through the brain. Glymphatic system activity increases during sleep or ketamine/xylazine (K/X) anesthesia, yet the mechanism(s) facilitating CSF influx are poorly understood. Here, we correlated influx of a CSF tracer into the brain with electroencephalogram (EEG) power, heart rate, blood pressure, and respiratory rate in wild-type mice under six different anesthesia regimens. We found that glymphatic CSF tracer influx was highest under K/X followed by isoflurane (ISO) supplemented with dexmedetomidine and pentobarbital. Mice anesthetized with α-chloralose, Avertin, or ISO exhibited low CSF tracer influx. This is the first study to show that glymphatic influx correlates positively with cortical delta power in EEG recordings and negatively with beta power and heart rate.