Peptidergic signaling from clock neurons regulates reproductive dormancy in Drosophila melanogaster.

With the approach of winter, many insects switch to an alternative protective developmental program called diapause. Drosophila melanogaster females overwinter as adults by inducing a reproductive arrest that is characterized by inhibition of ovarian development at previtellogenic stages. The insulin producing cells (IPCs) are key regulators of this process, since they produce and release insulin-like peptides that act as diapause-antagonizing hormones. Here we show that in D. melanogaster two neuropeptides, Pigment Dispersing Factor (PDF) and short Neuropeptide F (sNPF) inhibit reproductive arrest, likely through modulation of the IPCs. In particular, genetic manipulations of the PDF-expressing neurons, which include the sNPF-producing small ventral Lateral Neurons (s-LN(v)s), modulated the levels of reproductive dormancy, suggesting the involvement of both neuropeptides. We expressed a genetically encoded cAMP sensor in the IPCs and challenged brain explants with synthetic PDF and sNPF. Bath applications of both neuropeptides increased cAMP levels in the IPCs, even more so when they were applied together, suggesting a synergistic effect. Bath application of sNPF additionally increased Ca2+ levels in the IPCs. Our results indicate that PDF and sNPF inhibit reproductive dormancy by maintaining the IPCs in an active state. Author summary Diapause is a hormonally mediated process that allows insects to predict and respond to unfavourable conditions by altering their metabolism and behavior to resist the oncoming environmental challenges. In Drosophila melanogaster females a protective state of reproductive dormancy is induced by lower temperatures and shorter photoperiods that mimic the approach of winter. By genetically manipulating the circadian pacemaker s-LN(v)s cells, which express two neuropeptides, Pigment dispersing factor (PDF) and short Neuropeptide F (sNPF), we were able to modulate levels of gonadal arrest. PDF and sNPF appear to act as antagonists to dormancy, as do the Drosophila insulin-like peptides (dILPs) that are expressed in the insulin producing cells (IPCs). Indeed, we observe that the axonal projections from the s-LNvs appear to overlap with those from the IPCs implying that the clock cells signal to the IPCs. We confirm this possible communication by applying the two synthetic peptides to the IPCs and detecting a response in the IPC signal transduction pathway. We conclude that the clock neurons activate the IPCs via PDF and sNPF, which in turn release the dILPs, antagonise dormancy and lead to reproductive growth, thereby uncovering a neurogenetic circadian-overwintering axis.