Activity of RH-0345 on Ecdysteroid Production and Cuticle Secretion in Tenebrio molitor Pupae In Vivo and In Vitro

Abstract

The activity of RH-0345 (halofenozide), a novel benzoylhydrazine analogue, was investigated on ecdysteroid production, cuticle secretion, and hemolymph protein concentrations in Tenebrio molitor pupae. The compound was applied topically (5 and 10 μg/pupa) on newly ecdysed pupae or added to the culture medium (1 and 10 μM) of sternal integuments explanted from newly ecdysed pupae. Enzyme immunoassay measurements of ecdysteroid titers in the hemolymph revealed that RH-0345 applied topically on newly ecdysed pupae had no effect on the peak position of ecdysteroids in the hemolymph, but it caused a significant increase of the peak values. With in vitro cultured integument explants, RH-0345 caused a significant increase of the amounts of ecdysteroids released into the culture medium. Further, integuments were cultured in medium containing RH-0345 and/or 20-hydroxyecdysone (20E) and the effects on apolysis and cuticle growth were tested. In the presence of 20E alone, the epidermis was able to secrete a new cuticle under the apolyzed pupal cuticle. Similarly, integument explants from RH-0345-treated series cultured without 20E underwent apolysis and then secreted a new cuticle. The thickness of such a newly induced cuticle did not change as a function of the concentration of RH-0345 nor of the incubation period. In contrast, integument explants from RH-0345-treated series cultured in the presence of 20E secreted a new significantly thicker cuticle. The observed differences in cuticle thickness could be explained by differences in the protein content.