Abstract
The nanoscale organization of the F-actin cytoskeleton in neurons comprises membrane-associated periodical rings, bundles, and longitudinal fibers. The F-actin rings have been observed predominantly in axons but only sporadically in dendrites, where fluorescence nanoscopy reveals various patterns of F-actin arranged in mixed patches. These complex dendritic F-actin patterns pose a challenge for investigating quantitatively their regulatory mechanisms. We developed here a weakly supervised deep learning segmentation approach of fluorescence nanoscopy images of F-actin in cultured hippocampal neurons. This approach enabled the quantitative assessment of F-actin remodeling, revealing the disappearance of the rings during neuronal activity in dendrites, but not in axons. The dendritic F-actin cytoskeleton of activated neurons remodeled into longitudinal fibers. We show that this activity-dependent remodeling involves text {Ca}^{2+} and NMDA receptor-dependent mechanisms. This highly dynamic restructuring of dendritic F-actin based submembrane lattice into longitudinal fibers may serve to support activity-dependent membrane remodeling, protein trafficking and neuronal plasticity.
Highlights
The nanoscale organization of the F-actin cytoskeleton in neurons comprises membrane-associated periodical rings, bundles, and longitudinal fibers
We observed complex and diverse patterns of fluorescence inside the neuronal processes that were only distinguishable with super-resolution microscopy (Fig. 1)
While it was known that the dendritic lattice is distributed in a more patchy fashion, we show here that this difference is likely due, at least partially, to a distinct response to neuronal activity, which destabilizes the lattice only in dendrites and somata (Fig. 8)
Summary
The nanoscale organization of the F-actin cytoskeleton in neurons comprises membrane-associated periodical rings, bundles, and longitudinal fibers. The F-actin rings have been observed predominantly in axons but only sporadically in dendrites, where fluorescence nanoscopy reveals various patterns of F-actin arranged in mixed patches These complex dendritic F-actin patterns pose a challenge for investigating quantitatively their regulatory mechanisms. We developed here a weakly supervised deep learning segmentation approach of fluorescence nanoscopy images of F-actin in cultured hippocampal neurons This approach enabled the quantitative assessment of F-actin remodeling, revealing the disappearance of the rings during neuronal activity in dendrites, but not in axons. One of the hallmark discoveries made possible by fluorescence nanoscopy methods is the existence of a periodical lattice of F-actin, spectrin, and associated proteins under the surface membrane of neuronal processes. One feature that has not been controlled or assessed is the level of electrical or synaptic activity
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