A. Similar g‐ratios in callosal axons of male and female groups. To quantify effects of sex hormones on normal myelination, demyelination and remyelination, g‐ratios (diameter of axon/diameter of axon + myelin sheath) were measured. Electron micrographs of CC axons from normal myelination condition of gonadally intact females (ai) and males (aii) are shown. Calculated g‐ratio of intact male and female from NM, DM and RM groups show a significant increase in g‐ratio during the demyelination phase and a significant recovery during the remyelinating phase (b). B. Effect of OVX and estradiol (E2) replacement on callosal axon myelination. The effect of OVX and estradiol add‐back is represented by electron micrographs of CC axons from normal myelination groups. OVX significantly increased the number of thinly myelinated and non‐myelinated axons as compared to gonadally intact groups (ai,ii). OVX animals implanted with estradiol significantly increased the number of myelinated axons (bi,ii). Calculated g‐ratio of OVX alone and OVX + E2 callosal axons female from NM, DM and RM, group show a significant increase in g‐ratio during the demyelination phase and a significant recovery during the remyelination phase (c). Callosal axons from OVX + E2 groups during normal myelination and remyelination showed a significant decrease in g‐ratios as compared to OVX + placebo. C. Effect of castration and hormone replacement on callosal axon myelination. Castration also induced an increase in unmyelinated axons (ai,ii). Dihydrotestosterone (DHT), testosterone (T) and E2 pellets implanted in castrated males showed variable effects on recovery of axon myelination (a–d). There were less unmyelinated axons in castrated groups treated with either T or E2, but not DHT, which were similar to castrated animals implanted with placebo. Quantification of myelinated and non‐myelinated axons during cuprizone diet‐induced demyelination and subsequent remyelination showed a significant difference between groups. T and E2 implanted animals had more myelinated axons compared to placebo‐ and DHT‐implanted groups (t‐test *P < 0.05; n = 3, ANOVA; #P < 0.5; colors represent significance for NM, DM or RM. Images are at taken at 4800× (i) and 7200× (ii) and the scale bar is 1 μm.