The human gut microbiota undergoes compositional changes during pregnancy, and these bacterial changes have been shown to influence maternal metabolic responses; however, how this occurs is unclear. Part of these maternal metabolic changes include enhanced glucose stimulated insulin secretion (GSIS) and an expansion of pancreatic β cell mass (BCM). Providing a potential understanding of how the gut microbiota mediates this maternal response, we have new data suggesting a relationship may exist during pregnancy through the β cell-expressed free fatty acid receptor-2 (Ffar2), where this receptor is activated by nutrients derived from the gut microbiota, short chain fatty acids (SCFAs). Exploring this potential relationship, we have observed the following; 1) Ffar2 expression is altered in mouse islets by gestational insulin resistance, which suggests FFA2 may help in β cell adaption during pregnancy, 2) mice with a null mutation of FFA2 (Ffar2-/-) exhibit impaired glucose tolerance, lower insulin levels and diminished BCM expansion during pregnancy, 3) SCFAs, the primary ligands for FFAR2, are altered during pregnancy within the gut and blood of mice, and finally 4) the bacterial species in the mouse gut microbiota are influenced during pregnancy, as observed for humans. Because of these compelling data, our goal in this proposal is to dissect and test how the gut microbiota, SCFAs, and Ffar2 on β-cells are contributing to gestational glucose homeostasis during pregnancy. First, through a novel mouse model created by our group, a β cell specific knockout of Ffar2, we will explore how β cell expressed Ffar2 mediates GSIS and BCM changes during pregnancy. Next, we will identify the key fermentative gut bacterial taxa and the metagenomic profiles of these taxa that contribute to SCFA level changes during pregnancy. And finally, we will test the collective relationship between gut microbiota, SCFAs, and β cell expressed Ffar2 by modulating the gut microbiota through germ-free conditions, testing the influence of the gut microbiota on glucose homeostasis in our β cell specific knockout of Ffar2. If this novel relationship exists during pregnancy, we will reveal a new paradigm in gestational glucose metabolism, and provide new insight into how the gut microbiota influences host metabolism. !