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A major criticism is that the technique of high-pressure freezing only handles very small volumes at most 200 micrometers thick, and therefore, the tissue being from a mouse brain, a significant amount of injury to neuronal arbours was caused to generate such small samples.
Three kinds of samples were used:
(1) Cell culture neurons, which have their own problems and can't be considered authoritative on neuronal morphology.
(2) Hippocampal slices, which do recover from sectioning when in the right culture medium but only to some extent. Most neurons exist as fragments in the slice. Artifacts in morphologies are expected.
(3) Acutely extracted brain bits can't be immediately frozen; even a second is enough for neurons to fire and osmolarity to shape neuronal morphologies away from the natural state.
In summary: while surely neurons in their natural state don't look like those in textbooks, since all sample preparations suffer from artifacts, I am not convinced that this study resolves the issue. Try to freeze a small animal ā like it's been done for C. elegans. Do these peculiar axon morphologies exist in the HFP'ed worm?
The authors themselves admit that:
"treatments that disrupt these parameters like hyper- or hypo-tonic solutions, cholesterol removal, and non-muscle myosin II inhibition all alter the degree of axon pearling" ā and all of these come into play during sample preparation.
Preprint: https://www.biorxiv.org/content/10.1101/2023.07.20.549958v1.full
As published: https://www.nature.com/articles/s41593-024-01813-1
I wish the reviews were published. Andreas Prokop, a neuroscientist working on microtubules in neurons, was involved, which is reassuring.
#neuroscience #morphology #neurons #pearling #axons
