Basic NeuroscienceImmunohistochemical study of mouse sciatic nerves under various stretching conditions with “in vivo cryotechnique”

- “In vivo cryotechnique” (IVCT) revealed dynamic structure of sciatic nerve fibers.
- By mechanical stretching, sciatic nerve fibers showed a beaded appearance.
- Structural changes were related to sites of Schmidt-Lanterman incisures.
- Albumin was immunolocalized in nerve fibers, reflecting their living state.

BackgroundIn living animal bodies, some morphological changes of nerve fibers will probably occur when peripheral nerves are stretched or not stretched during various joint exercises. We aimed to capture the dynamic structures of nerves under various stretching conditions and to keep soluble serum proteins in their tissue sections.New methodMorphological changes of stretched or non-stretched sciatic nerve fibers were examined with “in vivo cryotechnique” (IVCT). Fibers were directly frozen with liquid isopentane-propane cryogen (−193 °C). Immunolocalizations of protein 4.1G and albumin were also examined in the fibers.ResultsThe structures of IVCT-prepared sciatic nerves under the stretched condition showed a beaded appearance. By immunostaining for membrane skeletal protein 4.1G, Schmidt-Lanterman incisures (SLIs) were clearly identified, and the heights of their circular truncated cones were increased at narrow sites of the nerve fibers under the stretched condition, compared to those of non-stretched nerve fibers. Albumin was immunolocalized in blood vessels and also along endoneurium including regions near the node of Ranvier.Comparison with existing methodsWith the conventional perfusion-fixation method (PF), it was difficult to keep stable postures of living mouse limbs for tissue preparation. In nerve fibers after PF, the structures of SLI were easily modified, and albumin was heterogeneously immunolocalized due to diffusion artifacts.ConclusionsIVCT revealed (1) the structures of peripheral nerve fibers under dynamically different conditions, indicating that the morphological changes of SLIs play a functional role as a bumper structure against mechanical forces, and (2) accurate immunolocalization of serum albumin in the sciatic nerve fibers.