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

Abstract

In clonal b-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 Ca V 1.2 exceeded that of Ca V 1.3 and Ca V 2.3 two-fold. Insulin secretion, which rose tenfold in response to 16 . 7 mM glucose, was completely abolished by 5 mM isradipine that blocks Ca V 1.2 and Ca V 1.3. Similarly, the increase in intracellular calcium in response to 15 mM glucose was decreased in the presence of 5 mM isradipine, and the frequency of calcium spikes was decreased to the level seen at 2 . 8 mM glucose. By contrast, inhibition of Ca V 2.3 with 100 nM SNX-482 did not significantly affect insulin secretion or intracellular calcium. Using RNA interference, Ca V 1.2 mRNA and protein levels were knocked down by w65% and w34% 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 mM isradipine. Calcium influx underlying this residual insulin secretion could result from persisting Ca V 1.2 expression in transfected cells since knock-down of Ca V 1.3 did not affect glucose-stimulated insulin secretion. In summary, our results suggest that Ca V 1.2 is critical for insulin secretion in INS-1 832/13 cells.

Comparative identification of Ca2+channel expression in INS-1 and rat pancreatic β cells

“…The L-type α1C subunit expression exceeded that of α1D 10-fold in INS-1 cells, but only 2-fold in the β cells. These results were similar to recent research in INS-1 832/12 cells which confirmed that α1C subunit expression exceeded that of the α1D subunit by twofold and that the α1C subunit had a critical role in insulin secretion [18] . The L-type α1C subunit performs a special function in the first phase of insulin secretion and glucose tolerance.…”

“…In the present study, L-type α1C subunits were dominant among all subunit expressions, which was in accordance with its important biological function in both INS-1 and rat β cells. The L-type α1D subunit's roles in insulin secretion or proliferation were not confirmed until recently [18,[20][21][22] . The expression of the non-voltage-operated Ca 2+ channel subunits Ryr2, TRPC1, TRPC4α, TRPC4β, and IP3R1 in rat β cells exceeded those in INS-1 by 10-fold and TRPM2 expression was not significantly different between the β cells and INS-1.…”

The human L-type calcium channel Cav1.3 regulates insulin release and polymorphisms in CACNA1D associate with type 2 diabetes

“…We also found that Ca v 1.3 knockdown reduced insulin release in INS-1 832/13 cells. A previous study suggested that Ca v 1.2 knockdown diminished GSIS [46] but as we did not test siRNA against Ca v 1.2, it may well be that both channels are crucial for appropriate insulin release in these cells.…”

Tight Coupling between Glucose and Mitochondrial Metabolism in Clonal β-Cells Is Required for Robust Insulin Secretion

“…However, neither expression of critical calcium channels nor insulin content is different in the two subclones (11). Furthermore, Newgard and co-workers (13) showed that glucose responsiveness in different INS-1 subclones correlates with an increased flux in an anaplerotic pathway, termed pyruvate cycling.…”

“…The 832/2 and 832/13 subclones express similar levels of Ca(V)1.2, Ca(V)1.3, and Ca(V)2.3 Ca 2ϩ channels, which are involved in GSIS in ␤-cells. This suggests that altered calcium channel activity is not responsible for discrepant insulin secretion responses (11). Furthermore, these clones are characterized by similar insulin content (12).…”