Effect of ecdysterone on RNA and DNA synthesis in the midgut of Bombyx mori was studied. It was found that ecdysterone stimulated in vivo RNA and DNA synthesis in the midgut of the I-day-old larva of the fifth instar. At first, stimulation of incorporation of ^3H-uridine into RNA occurred at the 6th hour after ecdysterone injection and continued from then. On the other hand, stimulation of incorporation of ^3H-thymidine into DNA occurred from the 18th hour after ecdysterone injection and silver grains were observed only in the nuclei of generative cells in autoradiography, and at the 48th hour after ecdysterone injection, new epithelium is formed as generative cell divided and multiplied. This indicates that RNA synthesis occurs in the nuclei of generative cell in the midgut of the larvae injected with ecdysterone, suggesting that ecdysterone stimulates first RNA synthesis for DNA synthesis. It may be understood that the action of ecdysterone on the midgut of the fifth instar larva as follow, ecdysterone acts on the generative cells which exists in the undifferentiated cells in the midgut tissue, induces the stimulation of RNA synthesis in the cell, the synthesized RNA induces it to synthesize DNA and subsequently the cell divides, multiplies and forms the new pupal epithelium. In order to examine the primary action of ecdysterone, one should try to use in vitro RNA synthesis in isolated nuclei. The isolated nuclei had the ability of RNA synthesizing activity. RNA synthesis elevated in the isolated midgut nuclei from 24th hour after ecdysterone injection. The result is consistent with similar effect observed with ecdysterone in the midgut nuclei in vivo RNA synthesis. The rate of RNA synthesis in the isolated midgut nuclei from various time after ecdysterone injection was examined. RNA synthesis enhanced greater in the isolated nuclei at 48th hour after ecdysterone injection than that at 1 hour. The rate of RNA synthesis in the isolated nuclei at 48th hour after ecdysterone injection increased linearly with incubation time, whereas the rate of RNA synthesis in the isolated nuclei after ecdysterone injection is lower than at the 48th hour and continued at lower level until 120 minutes after incubation. The result indicates that a physiological change, such as RNA synthesis elevated in the nuclei of the midgut, is developed by the action of ecdysterone. The direct effect of ecdysterone on RNA synthesis in the isolated nuclei was examined, by means of the nuclei isolated from the midgut of the 1-, 3- and 6-day-old larvae and incubated with ecdysterone. Each of the larvae were not injected with ecdysterone. There was little stimulation effect of ecdysterone in the isolated nuclei from the 1 and 3-day-old larvae, so-called the stage of feeding. On the other hand, in the isolated nuclei from the 6-day-old larvae, the stage of larval maturation, stimulation effect of ecdysterone on RNA synthesis increased by 30 per cent. It was found that there is a degree of responsiveness of nuclei from different stages to ecdysterone, and the nuclei in the midgut of the matured larva have more responsiveness to ecdystrone that that of the feeding larva.