CaV1.2 rather than CaV1.3 is coupled to glucose-stimulated insulin secretion in INS-1 832/13 cells.
In clonal beta-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 microM 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 microM 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 approximately 65% and approximately 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 microM 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.
Nitert
Marloes Dekker
MD
Molecular Metabolism Islet Cell Exocytosis, Department of Clinical Science, Clinical Research Center, SE-205 02 Malmö, Sweden Lund University Diabetes Center, Malmö, Sweden. marloes.dekker_nitert@med.lu.se
Nagorny
Cecilia L F
CL
Wendt
Anna
A
Eliasson
Lena
L
Mulder
Hindrik
H
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism / Q000502:physiology*; D020746:Calcium Channels, L-Type / Q000378:metabolism / Q000502:physiology*; D045744:Cell Line, Tumor; D002999:Clone Cells; D005947:Glucose / Q000502:physiology*; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism*; D007340:Insulinoma / Q000378:metabolism / Q000473:pathology; D017275:Isradipine / Q000494:pharmacology; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D051381:Rats
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D007328:Insulin; C419385:L-type calcium channel alpha(1C); C093712:Cacna1d protein, rat; D005947:Glucose; D017275:Isradipine