Many protocols for obtaining acute brain slices for electrophysiology involve cutting the brain in an ice-cold solution, then transferring the brain slices into a solution pre-warmed to near-physiological temperatures (up to 37 °C) for, e.g., 30 minutes, and then storing them at room temperature during the experiment. What is the purpose of the 30-minute warming step?
I've read that it kills unhealthy cells when using hyperosmolar sucrose-based solutions, but this step is also included when using isoosmolar solutions, including the standard recording ACSF.
Hi Alexey Galashin
The ice-cold aCSF freezes the cells and stops (partially) the metabolism of the cells to minimize the mechanical stress during the sectioning. After that you equilibrate your slices (1 hour) in physiological aCSF and you keep them there for the duration of your experiments. By doing so, you "trick" the slices into "believing" they are in a functioning environment. Your slices, when submerged in oxygenated physiological aCSF can be kept alive and properly functioning for up to 8 hours.
I hope this helps
Best
Dear Alexey,
In addition to Demos's answer, the incubation step rapidly kills and removes the damaged cells at the cut surface. These damaged cells cannot keep their integrity at physiological temperature and explode. The debris are washed away. This cleans the slice surface and allows you to better visualize the cells. By quickly getting rid of the damaged cells, it may reduce excitotoxicity and prolong the health of the tissue.
For adult brain slices the use of ice-cold slicing aCSF was found to be detrimental for the slice condition (see Article Acute Brain Slice Methods for Adult and Aging Animals: Appli...
andArticle Physiological temperature during brain slicing enhances the ...
), but they still use this step at physiological temperature right after slicing.
Good luck!