Gut microbiota as a modulator of circadian neural development in the honey bee model.

Abstract

In the past decade, it has become clear that gut microbes not only regulate digestion-related processes but also play an important role in the development and function of the central nervous system. Although gut microbiota are known to influence brain development and behavior, the mechanisms by which they affect specific neural circuits are not yet fully understood. We hypothesized that if gut microbiota is important for neural circuit development, then they may also influence the development of circadian circuits and the onset of behavioral circadian rhythm. In this thesis, we examined the role of gut microbiota in the onset of behavioral circadian rhythms and the maturation of the circadian clock mechanism using the honey bee as a model organism, which shows post-emergence development of circadian rhythms and clock system maturation similar to humans. First, we tested how gut microbiota affect the onset of behavioral circadian rhythms by applying three microbiota manipulations: antibiotic treatment, nurse bee interaction, and brood cap removal, and we measured locomotor activity under constant conditions. Our behavioral results show that depletion of gut microbiota reduced the percentage of bees that developed rhythmic behavior compared to controls, whereas microbial transfer from nurse bees did not produce a consistent effect. Next, we examined how gut microbiota influence neurons expressing the major circadian neuropeptide pigment-dispersing factor (PDF). Our results show that gut microbiota dysbiosis impairs the maturation of PDF-expressing neurons. Finally, to identify molecular pathways involved in circadian circuit development, We first performed a preliminary RNA-seq analysis on brain tissue and identified one notable candidate gene, Insulin-like Growth Factor Binding Protein Acid Labile Subunit (IGFALS), which is involved in insulin/IGF signaling. We then performed absolute-quantification qPCR to examine how antibiotic treatment affects IGFALS expression during early developmental stages. We found that bees exposed to antibiotic treatment had higher IGFALS copy numbers at early ages.<br /> <br /> Our model suggests that gut microbiota dysbiosis disrupts microbial signaling and increases IGFALS levels, which may retain IGF ligands in circulation and reduce their availability to bind receptors on clock neurons. This reduced IGF signaling may impair the maturation of clock neurons and, in turn, delay or reduce the development of behavioral circadian rhythms.