CaV1.2 rather than CaV1.3 is coupled to glucose-stimulated insulin secretion in INS-1 832/13 cells

Abstract

In clonal β-cell lines and islets from different species, a variety of calcium channels are coupled to glucose-stimulated insulin secretion. The aim of this study was to identify the voltage-gated calcium channels that control insulin secretion in insulinoma (INS)-1 832/13 cells. The mRNA level of CaV1.2 exceeded that of CaV1.3 and CaV2.3 two-fold. Insulin secretion, which rose tenfold in response to 16.7 mM glucose, was completely abolished by 5 μM isradipine that blocks CaV1.2 and CaV1.3. Similarly, the increase in intracellular calcium in response to 15 mM glucose was decreased in the presence of 5 μM isradipine, and the frequency of calcium spikes was decreased to the level seen at 2.8 mM glucose. By contrast, inhibition of CaV2.3 with 100 nM SNX-482 did not significantly affect insulin secretion or intracellular calcium. Using RNA interference, CaV1.2 mRNA and protein levels were knocked down by ∼65% and ∼34% respectively, which reduced insulin secretion in response to 16.7 mM glucose by 50%. Similar reductions in calcium currents and cell capacitance were seen in standard whole-cell patch-clamp experiments. The remaining secretion of insulin could be reduced to the basal level by 5 μM isradipine. Calcium influx underlying this residual insulin secretion could result from persisting CaV1.2 expression in transfected cells since knock-down of CaV1.3 did not affect glucose-stimulated insulin secretion. In summary, our results suggest that CaV1.2 is critical for insulin secretion in INS-1 832/13 cells.