Detection of insulin granule exocytosis by an electrophysiology method with high temporal resolution reveals enlarged insulin granule pool in BIG3-knockout mice

We recently identified BIG3 as a negative regulator of insulin granule biogenesis and reported increased insulin secretion in BIG3-knockout (BKO) mice. To pinpoint the site of action for BIG3, we investigated whether BIG3 regulates quantal insulin granule exocytosis. We established an assay to detect insulin granule exocytosis by recording ATP-elicited currents at high temporal resolution by patch clamp. Similarly to insulin, ATP release was increased in BKO β-cells. Although the frequency of insulin granule exocytosis was increased in BKO β-cells, quantal size or release kinetics remained unchanged. Electron microscopy studies showed that the number of insulin granules was increased by >60% in BKO β-cells. However, the number of morphologically docked granules was unaltered. The number of insulin granules having significant distances away from plasma membrane was greatly increased in BKO β-cells. Thus, BIG3 negatively regulates insulin granule exocytosis by restricting insulin granule biogenesis without the release kinetics of individual granules at the final exocytotic steps being affected. Depletion of BIG3 leads to an enlarged releasable pool of insulin granules, which accounts for increased release frequency and consequently increased insulin secretion