Stimulation of ecdysteroidogenesis by small prothoracicotropic hormone: role of calcium

Insect prothoracic glands are regulated by neuropeptide prothoracicotropic hormones (PTTH). In Manduca sexta PTTH exists as two size variants, big PTTH (approximately 25.5 kDa) and small PTTH (approximately 7 kDa). Previous studies indicate that both size variants employ cAMP as a second messenger and that stimulation of ecdysteroid secretion by big PTTH is Ca(2+)-dependent. In the present study, experiments were performed to assess the role of Ca2+ in small PTTH-stimulated ecdysteroid secretion by prothoracic glands from fifth instar larvae. Basal ecdysteroid secretion was not affected by Ca2+ channel blockers (verapamil or lanthanum) or by omission of Ca2+ from the incubation medium. Treatment of glands with a Ca2+ ionophore (A23187 or ionomycin) produced a concentration-dependent stimulation of ecdysteroid secretion. Stimulation of ecdysteroid secretion by small PTTH was suppressed (1) by Ca2+ channel blockers and (2) in Ca(2+)-free medium. A cAMP analog (Sp-cAMPS) stimulated ecdysteroid secretion in the presence of a Ca2+ channel blocker (verapamil) and in Ca(2+)-free incubation medium, and ionophore-induced ecdysteroid secretion appeared to be suppressed by a cAMP antagonist (Rp-cAMPS). The combined results indicate that basal ecdysteroid secretion is not dependent on external Ca2+, and suggest that small PTTH-stimulated ecdysteroid secretion is mediated by an influx of Ca2+ that precedes cAMP formation.