Abstract
L-Type Ca2+ channel blockers inhibit glucose and KCl-stimulated insulin secretion by pancreatic β cells. However, the role of the two distinct L-type channels expressed by β 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 recordings 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 transfected 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 completely inhibited and KCl-stimulated secretion was substantially resistant to inhibition by DHPs, but sensitive to ω-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.
Footnotes
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This work was supported by Grant 1119990378 from the American Diabetes Association (to G.H.H.).
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DOI: 10.1124/jpet.102.046334
- Abbreviations:
- VDCC
- voltage-dependent calcium channel
- DHP
- dihydropyridine
- DHPi
- dihyrdropyridine-insensitive
- SNARE
- solubleN-ethylmaleimide-sensitive factor attachment protein receptor
- GFP
- green fluorescent protein
- RT-PCR
- reverse transcription-polymerase chain reaction
- EGFP
- enhanced green fluorescent protein
- PCR
- polymerase chain reaction
- bp
- base pair(s)
- KRBH
- Krebs-Ringer-bicarbonate HEPES buffer
- IBa
- barium current
- Received October 30, 2002.
- Accepted December 30, 2002.
- The American Society for Pharmacology and Experimental Therapeutics
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