The effect of sex hormones on subendothelial connective tissue in rabbit arteries was sought, and striking morphological alterations were noted after prolonged treatment. Depot preparations of testosterone cypionate, 100 mg/week, or estradiol cypionate, 200 µg/week, were injected intramuscularly into intact adult New Zealand male rabbits for 8–16 weeks. The iliac arteries were prepared for electron-microscopic examination from five rabbits in each treatment group and in four normal male rabbits. Thin sections were prepared from three randomly selected cross-sectional tissue blocks for each animal. A minimum of 7 µm of vessel circumference was studied in each block. Selection of an area for electron-microscopic examination was random, except that those sites in which the internal elastic lamina showed fragmentation were discarded. Electron micrographs at a single final magnification were evaluated ‘blind’ by several observers for qualitative difference in subendothelial constituents. In the control group, the subendothelium contained moderate numbers of elastin-associated microfibrils (MF); all the rabbits given testosterone consistently showed significantly fewer MF than the control. In contrast, rabbits given estradiol all had significantly greater numbers of MF. Statistical analysis of variance showed these difference to be significant at p < 0.001.