Ultrastructural studies of the in vitro chitin and cuticle secretions in the wing pads of Locusta

Abstract

Cuticle deposition and chitin synthesis in the wing pads of 1-day-old fifth-instar larvae of Locusta were stimulated in vitro by 20-hydroxyecdysone and makisterone A. Stimulation was dose-dependent from 2 × 10−7 to 2 × 10−5 M. The response appeared after a 36- to 48-h lag period characterized by inhibition of the postecdysial activity of the previous cuticular cycle. The hormones stimulated deposition of the epicuticular layers, triggered uptake of the chitin precursor N-acetyl-D-glucosamine and biosynthesis of chitin, and initiated microtubular bundle orientation. Ecdysone has no effect on these processes. Ecdysone and 20-hydroxyecdysone do not act synergistically on chitin biosynthesis. These data confirm earlier observations on the differential effects of the major forms of ecdysteroids, ecdysone and 20-hydroxyecdysone, on the activity of epidermal cells of locusts. Chitin biosynthesis by 20-hydroxyecdysone stimulated cells requires the integrity of the microtubular system and the chitin–synthetase complex. It was inhibited by colcemid, a microtubular poison, and by diflubenzuron, an inhibitor of chitin synthetase. These drugs had no effect on apolysis and epicuticular secretion. The in vitro biosynthesis of chitin was also inhibited by azadirachtin, a cytotoxic drug.