Quantitative study of the non-circularity of myelinated peripheral nerve fibres in the cat

1. One hind limb of each of four cats was either chronically de-efferentated, or chronically de-afferentated, and perfused with buffered glutaraldehyde fixative. Up to three different muscle nerves were dissected from each limb, post-fixed in osmium tetroxide and embedded in Epon. Ultrathin transverse sections were mounted on Formvar-coated single-hole specimen grids so that all the fibres in each nerve could be examined individually by electron microscopy.2. Non-circularity was expressed as the ratio (ø): [Formula: see text] The degree of non-circularity of all the afferent axons, or all the efferent axons, in each muscle nerve was determined. The proportion of fibres cut through the paranodal region, or through the Schwann cell nucleus, was as expected for group I afferent and for alpha and gamma efferent fibres, but hardly any typical paranodal sections of group II or III afferent fibres were encountered which suggests that their paranodal arrangement differs from that of other groups. In a quantitative comparison of noncircularity in different functional groups, fibres cut through paranodes, Schwann cell nuclei or Schmidt-Lanterman clefts were rejected.3. All the gamma efferent fibres in one nerve were studied in a series of sections cut at 25 mum intervals. The degree of non-circularity was found to be relatively constant along the internode of most fibres when the values at paranodes, Schwann cell nuclei or Schmidt-Lanterman clefts were ignored.4. The value of ø varied widely from 1.0 (circular) to 0.5 or less from fibre to fibre within every functional group. However, the mean value of ø was less for gamma axons (0.68) than for alpha axons (0.78), and less for group III axons (0.79) than for axons in groups I and II (both 0.84). When the results for all the nerves were aggregated, these differences were statistically very highly significant, as was the difference in ø between group I and alpha fibres. If values of ø < 0.5 were rejected, the difference between the mean ø for group III and group II was then of doubtful significance whereas that between alpha and gamma fibres was still very highly significant.5. The external perimeter (S) of a non-circular fibre differs from pi times the diameter of a circle just enclosing the fibre (D). It is shown that S = 0.95 pi D for group I and II fibres, S = 0.90 piD for alpha and group III fibres, and S = 0.85 piD for gamma fibres.6. The myelin period, or interperiod repeat distance, varied from 14.1 to 15.6 nm in different cats, implying radial shrinkage of the myelin sheath from 15 to 23%. The myelin period in a particular cat was the same for several nerves, and the same for fibres in different functional groups.7. The possibility that repetitive firing of axons during fixation contributed to the varying degree of non-circularity is considered but rejected as unlikely.8. It is deduced that about 10% radial shrinkage of the myelin sheath, but little or no osmotic shrinkage of the axon, occurred during fixation and rinsing. Further radial shrinkage of about 8% in all components of the fibre probably occurred as a result of subsequent histological processing. It is concluded that the non-circularity of all axons, and the greater non-circularity of small axons, is unlikely to have been due to histological processing.9. It is concluded that axons are non-circular in vivo. The hypothesis that non-circularity allows axons to accommodate swelling during repetitive activity is discussed. Suggestions are made as to why gamma axons may be more non-circular than alpha or group III axons in an anaesthetized cat immediately prior to fixation, and why alpha axons may be more non-circular than axons in groups I and II.