Myelination determines the caliber of dorsal root ganglion neurons in culture

In order to understand the relationship of supporting cells to the differentiation of neurons in culture, we have used morphometry to study myelination of dorsal root ganglion (DRG) neurons by central or peripheral supporting cells. Dissociated DRG cultures from 15-day rat embryos, free of Schwann cells and fibroblasts, were prepared, and supporting cells were added from spinal cord or DRG; myelination commenced after 2 weeks. Control cultures received no supporting cells. At 7, 14, and 24 days, a total of 22 cultures were processed for electron microscopy. Three fascicles from defined points were sampled from each culture. In cultures containing glial cells, smaller fibers (p < 0.001) were myelinated (mean of median diameter, 1.13 ± 0.13 (SD) μm) than in cultures containing Schwann cells (1.67 ± 0.17 μm), although there was no difference (p > 0.1) in the degree of myelination expressed as number of myelin lamellae/fiber. A new finding concerned the relationship of axonal diameter to the presence or absence of myelinating cells. In control cultures without supporting cells or in areas where supporting cells were absent, the range of neurite diameter (0.05 to 1.25 μm) and the median diameter (mean of median, 0.24 ± 0.03 μm) were similar at different times (7, 14, and 24 days), demonstrating a stable population of neurite diameters throughout the period. In myelinated fascicles, a different distribution of neurite diameters was present. Myelinated neurites had a greater median diameter (measured to inner border of myelin) and a different range of fiber diameters compared to bare neurites. For Schwann cells, this range was 0.7 to 3.4 μm, and the mean of median diameters was 1.67 ± 0.17 μm; for glial cells, the range was 0.6 to 2.4 μm, and the mean of median diameters 1.13 ± 0.13 μm. Differences between myelinated and bare fibers were all highly significant (p < 0.001). The absence of the larger- diameter peak in fascicles containing only bare neurites suggests that myelination by Schwann cells or oligodendrocytes is necessary for the expression of axonal diameter greater than 1.25 μm in this tissue culture system.