In vivo monitoring of intracellular Ca(2+) dynamics in the pancreatic β-cells of zebrafish embryos

Assessing the response of pancreatic islet cells to glucose stimulation is important for understanding β-cell function. Zebrafish are a promising model for studies of metabolism in general, including stimulus-secretion coupling in the pancreas. We used transgenic zebrafish embryos expressing a genetically-encoded Ca(2+) sensor in pancreatic β-cells to monitor a key step in glucose induced insulin secretion; the elevations of intracellular [Ca(2+)](i). In vivo and ex vivo analyses of [Ca(2+)](i) demonstrate that β-cell responsiveness to glucose is well established in late embryogenesis and that embryonic β-cells also respond to free fatty acid and amino acid challenges. In vivo imaging of whole embryos further shows that indirect glucose administration, for example by yolk injection, results in a slow and asynchronous induction of β-cell [Ca(2+)](i) responses, while intravenous glucose injections cause immediate and islet-wide synchronized [Ca(2+)](i) fluctuations. Finally, we demonstrate that embryos with disrupted mutation of the Ca(V)1.2 channel gene cacna1c are hyperglycemic and that this phenotype is associated with glucose-independent [Ca(2+)](i) fluctuation in β-cells. The data reveal a novel central role of cacna1c in β-cell specific stimulus-secretion coupling in zebrafish and demonstrate that the novel approach we propose – to monitor the [Ca(2+)](i) dynamics in embryonic β-cells in vivo – will help to expand the understanding of β-cell physiological functions in healthy and diseased states.