Pupal development is elicited early in the last larval instar of the tobacco hornworm, Manduca sexta (Johannson), by a precise temporal and quantitative increase in the haemolymph titre of 20-hydroxyecdysone. This increase in the titre is referred to as the pupal commitment peak, and it occurs once the titre of juvenile hormone (JH) has dropped. If the haemolymph titre of JH remains elevated at this time due to topical application of the hormone or of its analogue ZR512, commitment is delayed or inhibited in a dose-dependent manner. This delay or inhibition is due to the curtailment of the commitment peak in the ecdysteroid titre, which results from a failure of the prothoracic glands (PG) to increase the synthesis/secretion of the premoulting hormone, ecdysone. Since the PG from ZR512- and JH 1-treated larvae are capable of being activated in vitro by the prothoracicotropic hormone (PTTH), the effect of JH on the PG does not involve suppression of gland sensitivity to PTTH. The locus of the JH effect was determined to be the brain-retrocerebral complexes (Br-CC-CA), on the basis of experiments which tested the effect of implanted Br-CC-CA from pre-commitment larvae treated with JH on the occurrence of pupal commitment in head-ligated larval hosts. The implanted, JH-treated Br-CC-CA exhibited a delayed release of PTTH, and the effect was at concentrations of JH that were physiological. These results argue that JH functions to control the time during the last larval instar when pupal commitment occurs by dictating when PTTH will undergo gated release.