Type 1 Diabetes (T1D) generally results from a poorly understood autoimmune process that leads to selective destruction of pancreatic beta cells. Interestingly, beta cell function is often not correlated with mass and declines very early in the disease before symptom onset, but the mechanisms driving this are not known. Islet beta cells existing in a functional hierarchy, with a small proportion of key beta cells exerting a disproportional effect on overall islet insulin release. These key beta cells, termed ‘hub’ and ‘leader’ cells are the sources of calcium oscillations and coordinate connectivity between other ‘follower’ beta cells, thereby governing the amplitude and duration of insulin secretory granule release. Previous work has also shown that hub and leader beta cells are a stable beta cell subtypes that are sensitive to metabolic and inflammatory stress in vitro, but whether loss or dysfunction of these beta cell subtypes are causally linked to overall beta cell functional decline and inter-individual heterogeneity in T1D has not been explored. In this study, we propose to test the hypothesis that hub and leader beta cells are lost early on in T1D, which would lead to extensive heterogeneity in islet insulitis and functional beta cell mass both between individuals at the same disease stage and between different disease stages in the same individual. To test these ideas, we will carry outlive cell imaging and functional studies on live human donor pancreas slices from individuals at different disease stages (no diabetes/control, early-late autoimmunity, recent onset T1D) using genetically targeted probes to track calcium dynamics. We will apply computational approaches to characterize beta cell functional hierarchy in these slices, as well as use of optical probes to photo-label hub and leader populations in situ for analysis of their behavior in the autoimmune slice milieu. We propose to further characterize the inter-relationships between hub and leader beta cells, beta cell stress pathways and islet insulitis using fixed pancreas sections. Finally, using the live slice in situ model, we will explore how small molecules to that selectively beta cell stress pathways (including senescnece and unfolded protein response)impacts on islet functional hierarchy and connectivity. Together these studies will take us closer to understanding the contributions of beta cell heterogeneity to T1D heterogeneity.