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The distribution of prothoracicotropic hormone in the pupal brain of Manduca sexta has been determined by an in vitro assay for prothoracic gland activation. Prothoracicotropic activity was observed in both the brain and retrocerebral complex, but predominantly in the dorsolateral regions of the protocerebrum. Of the two groups of neurosecretory cells present in this area of the brain, only the two lateral type III neurosecretory cells exhibited significant prothoracicotopic hormone activity. Further analysis revealed that the neurohormone was localized in only one of the two type III cells, suggesting that a single neurosecretory cell in each hemisphere is the source of the hormone at the stage examined (day 0). Prothoracicotropic hormone activity was detected in both the corpora allata and the corpora cardiaca, but the corpora allata contained 2 to 9 times the activity of the corpora cardiaca, depending on developmental stage. The significantly higher level of activity in the corpora allata suggests that they may be the neurohemal organs through which the prothoracicotropic hormone of Manduca is released.The role of the brain in the neuroendocrine control of insect molting was demonstated about 4 decades ago in a series of classical experiments utilizing ligation, decapitation, and implantation techniques (1). Since that time, various other approaches, both indirect (2-5) and direct (6), have confirmed the tenet that the brain produces a neurohormone that stimulates the prothoracic glands to synthesize the prohormone ecdysone (7), whose hydroxylation product ecdysterone then elicits the molt. Although the prothoracicotropic hormone (PTTH) is the primary effector of insect postembryonic development (8), neither the source of PTTH within the brain nor the site of its release into the hemolymph has been directly demonstrated.It is presumed that PTTH is the product of specific cerebral neurosecretory cells (NSC) (9). Studies attempting to localize PTTH by the implantation of different groups of NSC or injection of homogenates of portions of the brain have yielded various results among the several insect species investigated. For example, in the reduviid bug Rhodnius prolixus, and the aphid Megoura viciae, PTTH activity appears to be located in the medial neurosecretory cells (M-NSC) of the protocerebrum (10, 11), whereas in the sphingid Manduca sexta the lateral neurosecretory cells (L-NSC) of the pupal brain may be the source of the neurohormone (5). By contrast, in the saturniid Hyalophora cecropia, both M-NSC and L-NSC are thought to produce PTTH (12).Definitive identification of the NSC that produce PTTH has not been made, in part due to the nature of the bioassays used to assess hormone activity (6, 9). However, with the recent development of a specific and sensitive in vitro assay for PTTH (6), the localization of this neurohormone in specific cerebralThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in ac...