High temporal resolution, in vivo imaging for improved measurements of cerebrospinal fluid flow in mice*

The glymphatic system, a recently discovered pathway for flow of cerebrospinal fluid (CSF) in the brain, has been shown to play a key role in the clearance of toxic cellular waste from the brain's extracellular space. In vivo visualization of this system however is challenging due to lack of optical access through the opaque skull, the fragile nature of the brain environment, and the difficulty of experiments due to invasive surgeries. Several imaging techniques have been used to image this system in vivo such as transcranial fluorescence macroscopy, functional magnetic resonance imaging, and confocal microscopy; however, the coarse spatial and temporal resolutions that can be achieved with these techniques also limit our ability to draw conclusions about driving mechanisms of CSF flow. To overcome these limitations, prior studies (e.g., Mestre et al 2018) employed two-photon microscopy, where they were able to measure CSF velocities with imaging resolutions on the order of 1 µm at 30 Hz. Their work showed that arterial pulsations stemming from the cardiac cycle are a key driver of CSF flow, but the chosen imaging rate was still rather coarse for the mouse heart rate (~6 Hz). To build on their work, we imaged CSF flow at rates as high as 440 Hz. This drastic increase in temporal resolution enables rigorous quantification of how CSF pulses in synchrony with arterial pulsations and improved estimates of the wall shear stresses experienced by pial vessels and astrocytes due to glymphatic flow.