Tension Sensor Based on Fluorescence Resonance Energy Transfer Reveals Fiber Diameter-Dependent Mechanical Factors During Myelination.

Takeshi Shimizu,Hideji Murakoshi,Atsunori Hattori,Naoki Tajiri,Kota Ichino,Hidetoshi Matsumoto,Akimasa Ishida,Shinya Ueno,Hideki Hida

doi:10.3389/fncel.2021.685044

Abstract

Oligodendrocytes (OLs) form a myelin sheath around neuronal axons to increase conduction velocity of action potential. Although both large and small diameter axons are intermingled in the central nervous system (CNS), the number of myelin wrapping is related to the axon diameter, such that the ratio of the diameter of the axon to that of the entire myelinated-axon unit is optimal for each axon, which is required for exerting higher brain functions. This indicates there are unknown axon diameter-dependent factors that control myelination. We tried to investigate physical factors to clarify the mechanisms underlying axon diameter-dependent myelination. To visualize OL-generating forces during myelination, a tension sensor based on fluorescence resonance energy transfer (FRET) was used. Polystyrene nanofibers with varying diameters similar to neuronal axons were prepared to investigate biophysical factors regulating the OL-axon interactions. We found that higher tension was generated at OL processes contacting larger diameter fibers compared with smaller diameter fibers. Additionally, OLs formed longer focal adhesions (FAs) on larger diameter axons and shorter FAs on smaller diameter axons. These results suggest that OLs respond to the fiber diameter and activate mechanotransduction initiated at FAs, which controls their cytoskeletal organization and myelin formation. This study leads to the novel and interesting idea that physical factors are involved in myelin formation in response to axon diameter.

Highlights

In the central nervous system (CNS) white matter, large and small caliber axons are intermingled, and the diameter of myelin internodes is highly divergent (Weruaga-Prieto et al, 1996), especially in the spinal cord
Previous studies have reported that focal adhesions (FAs) act as mechanotransducers that transmit various signaling pathways (Geiger et al, 2009), which regulate cell morphogenesis and dynamics. These and our results indicate that physical factors are involved in myelin formation in response to axon caliber by activating mechanical signaling initiated at FAs
Since fluorescence resonance energy transfer (FRET) is known to shorten the fluorescence lifetime of the donor fluorophore (Murakoshi, 2021), we measured the fluorescence lifetime changes at fluorescent positive OL processes contacting nanofibers compared to the control area

Summary

Introduction

In the central nervous system (CNS) white matter, large and small caliber axons are intermingled, and the diameter of myelin internodes is highly divergent (Weruaga-Prieto et al, 1996), especially in the spinal cord. Optimization of the g-ratio is important for higher brain functions This indicates that myelin formation is tightly associated with the axon caliber, involving unknown diameter-dependent regulatory factors. Lee et al (2012) have previously reported that OL can ensheath a myelin membrane on artificial electrospinning nanofibers without living neurons These reports indicate that molecular signaling activated by functional proteins on the axonal surface is not required for initiation of myelination, but rather there are permissive axonal cues that initiate myelination (Lee et al, 2012). They investigated the effect of fiber diameter on myelination using varying nanofiber diameters (0.2–4.0 μm), revealing that larger diameter fibers (more than 0.4 μm) were preferentially ensheathed by OLs. the mechanisms underlying optimal myelination according to axon diameter have not been fully elucidated

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Conclusion