In the anterior silk glands (ASGs) of the silkworm, Bombyx mori, intracellular cAMP increases transiently to a very high level shortly after the hemolymph ecdysteroid peak in the prepupal period. In cultured ASGs obtained on the day of gut-purge, cAMP levels were increased by 20-hydroxy-ecdysone (20E), and this increase was enhanced by an inhibitor of phosphodiesterase, but was not affected by α-amanitin, indicating the 20E action may not be mediated via gene expression. The increase in cAMP occurred within 30 seconds of exposure to a physiological concentration of 20E (1 μM), and also by ponasterone A. Our findings indicate a nongenomic action of ecdysteroids in insects, which may be an additional mechanism by which this steroid hormone induces acute responses in tissues and cells.