Heterogeneity of glucose-stimulated insulin secretion (GSIS) in pancreatic islets is physiologically important but poorly understood. Here, we utilize mouse islets to determine how microtubules (MTs) affect secretion toward the vascular extracellular matrix at single cell and subcellular levels. Our data indicate that MT stability in the β-cell population is heterogenous, and that GSIS is suppressed in cells with highly stable MTs. Consistently, MT hyper-stabilization prevents, and MT depolymerization promotes the capacity of single β-cell for GSIS. Analysis of spatiotemporal patterns of secretion events shows that MT depolymerization activates otherwise dormant β-cells via initiation of secretion clusters (hot spots). MT depolymerization also enhances secretion from individual cells, introducing both additional clusters and scattered events. Interestingly, without MTs, the timing of clustered secretion is dysregulated, extending the first phase of GSIS and causing oversecretion. In contrast, glucose-induced Ca2+ influx was not affected by MT depolymerization yet required for secretion under these conditions, indicating that MT-dependent regulation of secretion hot spots acts in parallel with Ca2+ signaling. Our findings uncover a novel MT function in tuning insulin secretion hot spots, which leads to accurately measured and timed response to glucose stimuli and promotes functional β-cell heterogeneity.

37Heterogeneity of glucose-stimulated insulin secretion (GSIS) in pancreatic islets is 38 physiologically important but poorly understood. Here, we utilize whole mouse islets to 39 determine how microtubules affect secretion toward the vascular extracellular matrix. Our data 40indicate that microtubule stability in the β-cell population is heterogenous, and that cells with 41 more stable microtubules secrete less in response to a stimulus. Consistently, microtubule 42hyper-stabilization prevents, and microtubule depolymerization promotes β-cell activation. 43Analysis of spatiotemporal patterns of secretion events shows that microtubule 44 depolymerization activates otherwise dormant β-cells via initiation of secretion clusters (hot 45 spots). Microtubule depolymerization also enhances secretion from individual cells, introducing 46both additional clusters and scattered events. Interestingly, without microtubules, the timing of 47 clustered secretion is dysregulated, extending the first phase of GSIS. Our findings uncover a 48 novel microtubule function in tuning insulin secretion hot spots, which leads to accurately 49 measured and timed response to glucose stimuli and promotes functional β-cell heterogeneity. 50

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