Toward dynamic, anisotropic, high-resolution, and functional measurement in the brain extracellular space.

Diffusion of substances in the brain extracellular space (ECS) is important for extrasynaptic communication, extracellular ionic homeostasis, drug delivery, and metabolic waste clearance. However, substance diffusion is largely constrained by the geometry of brain ECS and the extracellular matrix. Investigating the diffusion properties of substances not only reveals the structural information of the brain ECS but also advances the understanding of intercellular signaling of brain cells. Among different techniques for substance diffusion measurement, the optical imaging method is sensitive and straightforward for measuring the dynamics and distribution of fluorescent molecules or sensors and has been used for molecular diffusion measurement in the brain. We mainly discuss recent advances of optical imaging-enabled measurements toward dynamic, anisotropic, high-resolution, and functional aspects of the brain ECS diffusion within the last 5 to 10 years. These developments are made possible by advanced imaging, such as light-sheet microscopy and single-particle tracking in tissue, and new fluorescent biosensors for neurotransmitters. We envision future efforts to map the ECS diffusivity across the brain under healthy and diseased conditions to guide the therapeutic delivery and better understand neurochemical transmissions that are relevant to physiological signaling and functions in brain circuits.