Volumetric Characterization of CNS Extracellular Space using Thick Undehydrated Hippocampal Tissue Cryosections
This study utilizes thick undehydrated hippocampal cryosections to characterize the CNS extracellular space, revealing that over 25% of the tissue volume consists of a porous network of narrow openings and larger vascular-adjacent voids that facilitate CSF flow and cell motility.
Original paper licensed under CC BY 4.0 (https://creativecommons.org/licenses/by/4.0/). This is an AI-generated explanation of a preprint that has not been peer-reviewed. It is not medical advice. Do not make health decisions based on this content. Read full disclaimer
Inside the brain, the space between cells is just as critical as the cells themselves. For decades, scientists have focused on the solid parts of the brain—the neurons that fire signals and the glial cells that support them. However, the fluid that fills the gaps between these cells, known as interstitial fluid, is equally vital. This fluid acts as a delivery system, bringing nutrients to cells and carrying away waste, while also allowing immune cells to patrol the tissue for damage. Without enough room for this fluid to move, the brain's delicate balance breaks down, potentially leading to disease. Understanding exactly how much space exists between cells, and what that space looks like, has been a difficult puzzle because standard methods for studying brain tissue often squeeze the fluid out, collapsing the very gaps researchers are trying to measure.
A new study by Jesus Trejos at St. John's University offers a fresh look at this hidden landscape by changing how the brain tissue is prepared. Instead of using the traditional method of drying out thin slices of brain tissue, which shrinks the gaps, the researcher examined thick, wet slices of the hippocampus—a region of the brain essential for memory. By keeping the tissue hydrated and cutting it into sections 60 microns thick, the team was able to see the brain's internal structure in a state much closer to how it exists in a living animal. This approach revealed that the brain is not a solid block of cells, but a highly porous structure filled with open channels and tunnels.
The researchers found that empty space makes up a significant portion of the brain's volume. In healthy tissue, roughly 25% of the space is not occupied by cells or solid structures. This empty space appears as a network of openings ranging from tiny 5-micron gaps to much larger voids up to 200 microns wide. These openings are not random holes; they often form circular or tunnel-like shapes that seem to connect different parts of the tissue. Some of the larger voids are found right next to blood vessels, suggesting they might serve as major pathways for fluid to enter or leave the brain tissue. The study also observed that these spaces are lined by cells, particularly microglia, which are the brain's immune sentinels. These cells appear to arrange themselves around the openings, perhaps to monitor the flow of fluid and detect any signs of trouble.
To confirm that these spaces were real and not just an illusion created by the preparation process, the team looked at what happened when the tissue was injured. When brain cells were damaged or died, the tissue collapsed. The empty spaces shrank or disappeared entirely, and the solid parts of the tissue pressed closer together. This collapse was visible under the microscope as a loss of the dark, open areas that represented the fluid-filled gaps. In contrast, healthy tissue maintained its open, porous structure. This observation proved that the spaces depend on living, functioning cells to hold their shape. If the cells die, the pathways for fluid flow close up, which could explain why fluid movement stops in damaged areas of the brain.
The study also explored how these spaces allow substances to move through the brain. When the researchers placed a drop of fluid on top of a thick slice of brain tissue, the fluid was able to travel all the way through to the bottom. This movement would be impossible if the tissue were solid or if the gaps between cells were too small. The fact that fluids could pass through the 60-micron-thick slices suggests that the brain is designed with a built-in plumbing system that allows for efficient circulation. This system is likely essential for the brain to maintain its steady state, ensuring that cells receive what they need and get rid of what they don't.
One of the most striking findings was the relationship between the shape of the cells and the spaces they create. Many brain cells have a specific structure where their extensions curve away from the main body, creating a natural gap. When many of these cells are arranged together, these individual gaps line up to form the larger tunnels and channels observed in the study. This structural arrangement suggests that the brain's architecture is naturally designed to facilitate the flow of fluid. The study proposes that this flow is not just a passive occurrence but a critical part of how the brain functions, supporting everything from the electrical signals that allow us to think to the immune responses that protect us from infection.
By using thick, undried sections of brain tissue, this research provides a clearer picture of the brain's internal geography than has been possible before. It challenges the old view of the brain as a dense, solid mass and replaces it with an image of a dynamic, porous sponge where fluid and cells coexist in a carefully balanced relationship. The findings suggest that the health of the brain depends not just on the cells themselves, but on the spaces between them remaining open and functional. If these spaces are compromised, whether by injury, disease, or cell death, the brain's ability to clean itself and communicate internally could be severely disrupted. This new understanding of the brain's volumetric layout opens the door to better ways of studying brain health and could eventually help scientists develop new treatments for conditions where fluid flow is impaired.
Drowning in papers in your field?
Get daily digests of the most novel papers matching your research keywords — with technical summaries, in your language.