Cav1.3 is preferentially coupled to glucose-stimulated insulin secretion in the pancreatic beta-cell line INS-1

L-Type Ca2 channel blockers inhibit glucose and KCl-stimu-lated insulin secretion by pancreatic cells. However, the role of the two distinct L-type channels expressed by cells, Cav1.2 and Cav1.3, in this process is not clear. Therefore, we stably transfected INS-1 cells with two mutant channel constructs, Cav1.2DHPi or Cav1.3 DHPi. Whole-cell patch-clamp record-ings demonstrated that both mutant channels are insensitive to dihydropyridines (DHPs), but are blocked by diltiazem. INS-1 cells expressing Cav1.3/DHPi maintained glucose- and KCl-stimulated insulin secretion in the presence of DHPs, whereas cells expressing Cav1.2/DHPi demonstrated DHP resistance to only KCl-induced secretion. INS-1 cells were also stably trans-fected with cDNAs encoding the intracellular loop between domains II and III of either Cav1.2 or Cav1.3 (Cav1.2/II-III or Cav1.3/II-III). Glucose- and KCl-stimulated insulin secretion in Cav1.2/II-III cells were not different from untransfected INS-1 cells. However, glucose-stimulated insulin secretion was com-pletely inhibited and KCl-stimulated secretion was substantially resistant to inhibition by DHPs, but sensitive to -agatoxin IVA in Cav1.3/II-III cells. Moreover, the L-type channel agonist FPL 64176 markedly enhanced KCl-stimulated secretion by Cav1.3/ II-III cells. Together, our results suggest that Ca2 influx via Cav1.3 is preferentially coupled to glucose-stimulated insulin secretion in INS-1 cells. Calcium influx through voltage-dependent Ca2 channels (VDCCs) plays a crucial role in insulin secretion from pan-creatic cells (Wollheim and Sharp, 1981). Glucose metabo-lism in cells causes an increase in ATP/ADP ratio, which closes ATP-dependent potassium channels (Rajan et al., 1990). The resulting membrane depolarization opens VD-CCs, and calcium influx triggers insulin secretion. Several distinct VDCCs have been detected in cells and insulin-secreting cell lines, including high-voltage-activated sub