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The functional organization of central motor circuits underlying feeding behaviors is not well understood. We have combined electrophysiological and genetic approaches to investigate the regulatory networks upstream of the motor program underlying food intake in the Drosophila larval central nervous system. We discovered that the serotonergic network of the CNS is able to set the motor rhythm frequency of pharyngeal pumping. Pharmacological experiments verified that modulation of the feeding motor pattern is based on the release of serotonin. Classical lesion and laser based cell ablation indicated that the serotonergic neurons in the subesophageal zone represent a redundant network for motor control of larval food intake.

Summary Steroid hormones play key roles in development, growth, and reproduction in various animal phyla [ 1 ]. The insect steroid hormone, ecdysteroid, coordinates growth and maturation, represented by molting and metamorphosis [ 2 ]. In Drosophila melanogaster, the prothoracicotropic hormone (PTTH)-producing neurons stimulate peak levels of ecdysteroid biosynthesis for maturation [ 3 ]. Additionally, recent studies on PTTH signaling indicated that basal levels of ecdysteroid negatively affect systemic growth prior to maturation [ 4 , 5 , 6 , 7 , 8 ]. However, it remains unclear how PTTH signaling is regulated for basal ecdysteroid biosynthesis. Here, we report that Corazonin (Crz)-producing neurons regulate basal ecdysteroid biosynthesis by affecting PTTH neurons. Crz belongs to gonadotropin-releasing hormone (GnRH) superfamily, implying an analogous role in growth and maturation [ 9 ]. Inhibition of Crz neuronal activity increased pupal size, whereas it hardly affected pupariation timing. This phenotype resulted from enhanced growth rate and a delay in ecdysteroid elevation during the mid-third instar larval (L3) stage. Interestingly, Crz receptor (CrzR) expression in PTTH neurons was higher during the mid- than the late-L3 stage. Silencing of CrzR in PTTH neurons increased pupal size, phenocopying the inhibition of Crz neuronal activity. When Crz neurons were optogenetically activated, a strong calcium response was observed in PTTH neurons during the mid-L3, but not the late-L3, stage. Furthermore, we found that octopamine neurons contact Crz neurons in the subesophageal zone (SEZ), transmitting signals for systemic growth. Together, our results suggest that the Crz-PTTH neuronal axis modulates ecdysteroid biosynthesis in response to octopamine, uncovering a regulatory neuroendocrine system in the developmental transition from growth to maturation.

Growth and maturation are coordinated by steroid hormone biosynthesis in various animal phyla. The insect steroid hormone, ecdysteroid, coordinates growth and maturation, represented by molting and metamorphosis. In Drosophila melanogaster, prothoracicotropic hormone (PTTH) neurons stimulate in generating peak levels of ecdysteroid to trigger maturation. In addition, recent studies have shed light on the role of PTTH signaling in basal ecdysteroid biosynthesis, which negatively affects systemic growth prior to maturation. However, it remains unclear how PTTH signaling is regulated for basal ecdysteroid biosynthesis. Here, we report that Corazonin (Crz)-producing neurons regulate basal ecdysteroid biosynthesis by affecting PTTH neurons. Inhibition of Crz neuronal activity increased pupal size, whereas it had little effect on pupariation timing. This phenotype resulted from enhanced growth and a delay in basal ecdysteroid elevation during the mid-third instar larval (L3) stage. Silencing of Crz in Crz neurons resulted in increased levels of the microRNA bantam, which represses basal ecdysteroid biosynthesis. Interestingly, Crz receptor (CrzR) expression in PTTH neurons was higher during the mid- than during the late-L3 stage. Silencing of CrzR in PTTH neurons increased pupal size, phenocopying the inhibition of Crz neuronal activity. When Crz neurons were optogenetically activated, a strong calcium response was observed in PTTH neurons in the mid-, but not the late-L3 stage. These data suggest that the Crz-PTTH neuronal axis modulates basal, but not peak ecdysteroid biosynthesis. Most significantly, this study uncovered a regulatory neuronal system affecting ecdysteroid biosynthesis in a developmental stage-specific manner.

The enteric nervous system is critical for coordinating diverse feeding-related behaviors and metabolism. We have characterized a cluster of four serotonergic neurons in Drosophila larval brain: cell bodies are located in the subesophageal ganglion (SOG) whose neuronal processes project into the enteric nervous system. Electrophysiological, calcium imaging and behavioral analyses indicate a functional role of these neurons in modulating foregut motility. We suggest that the axonal projections of this serotonergic cluster may be part of a brain–gut neural pathway that is functionally analogous to the vertebrate vagus nerve.