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Durham University

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Publication details for Dr Robert William Banks

Durbaba, R., Rudd, D., Hulliger, M. & Banks, R. W. (2012), Assessment of myelin thickness and axon circularity in a dorsal root of the mouse, Proceedings of the Physiological Society 27: Physiology 2012. Edinburgh, Physiological Society, PC261.

Author(s) from Durham

Abstract

Previous studies of peripheral nerves and ventral roots have shown that myelin thickness increases non-linearly with axon size and that axon circularity may be dependent on axon size (e.g. Arbuthnott et al. 1980; Biscoe et al. 1982; Ceballos et al. 1999). In this study, we present preliminary data relating myelin thickness and axon circularity to axon size for the central end of a dorsal root in a mouse cervical segment. A CD1 strain mouse was anaesthetized with sodium pentobarbitone (50 mg.kg-1 I.P.) and fixed by transcardial perfusion with a Karnovsky fixative. The left C5 dorsal root ganglion, along with the corresponding dorsal and ventral roots, was removed and placed in fresh fixative overnight at 4°C. The tissue was post-fixed with OsO4 prior to embedding in Araldite. Semithin serial sections (0.5 μm) were cut using an ultramicrotome and stained with 1% toluidine blue in 1% borax. A composite photomicrograph of the central end of the C5 dorsal root was constructed with a resolution of 10 pixels/μm and a magnification of x1000. Areas and perimeters of nerve fibres (axon plus myelin) and of their axons alone were determined using the freeware program Reconstruct (Boston University, Boston, MA, USA; see Fiala, 2005). Fibres were excluded from the analysis if they displayed internal folds, Schwann cell nuclei, Schmidt-Lantermann incisures or paranodal characteristics. From the area measures, axon (DA) and fibre (DF) diameters were computed for the equivalent circles. Myelin thickness was calculated as (DF - DA)/2. An index of circularity (IC) of the axon was computed by dividing the observed axonal area by the area of a circle with the same perimeter. Observations on 590 fibres, revealed that for DA≤6μm, myelin thickness increased linearly with a slope of 0.15. For DA>6μm, myelin thickness remained approximately constant (regression slope of -0.03), with a mean value of 1.17μm. Thus, myelin thickness initially increases with axon size prior to reaching a plateau. This conclusion is consistent with data from myelinated axons in peripheral nerves in the mouse (Ceballos et al. 1999). The IC has been reported to depend on axon size (Biscoe et al. 1982; Ceballos et al. 1999) but the results are conflicting. Biscoe et al. (1982) report higher values for smaller axons, whilst Ceballos et al. (1999) reported the converse. Our data for IC showed no clear tendency to vary with axon size, with an overall mean value of 0.78. Possible explanations for the differences between these sets of data might be that: (i) the observations of Biscoe et al. (1982) were based on lumbar ventral roots which would also include an autonomic component; (ii) our criteria for exclusion of axon profiles appear to be more stringent than those of Ceballos et al. (1999).