Muscarinic modulation of voltage-dependent Ca2+ channels in insulin- secreting HIT-T15 cells

Jeffrey A. Love, Neil W. Richards, Chung Owyang, David C. Dawson

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

Potentiation of insulin secretion from pancreatic β-cells by acetylcholine requires ongoing cyclic electrical activity initiated by other depolarizing secretagogues. Patch-clamp recordings in glucose-free solutions were made from the clonal β-cell line HIT-T15 to determine whether the muscarinic agonist bethanechol (BCh) modulated voltage-dependent Ca2+ channels independent of effects on membrane potential. Only high-threshold, dihydropyridine-sensitive (L-type) Ca2+ channels with a mean conductance of 26 pS were observed in cell-attached patches. BCh (100 μM) caused a two- to threefold increase in both fractional open time and mean current of single Ca2+ channels. These changes resulted from a 44% decrease in the longer of two apparent mean closed times and a 25% increase in the mean open time. Similar BCh-stimulated increases in macroscopic Ca2+ currents were recorded in whole cell, perforated-patch recordings. The role of protein kinase C (PKC) in the muscarinic activation of Ca2+ channels was tested using a variety of PKC activators and inhibitors. Acute application of either the active phorbol ester phorbol 12-myristate 13-acetate (PMA) or the membrane- permeable diacylglycerol analog 1,2-didecanoyl-rac-glycerol mimicked the effects of BCh, whereas an inactive phorbol (4α) had no effect. Depletion of PKC activity by chronic exposure to PMA or acute application of the PKC inhibitor staurosporine greatly reduced or abolished muscarinic activation of Ca2+ channels. These results are consistent with muscarinic activation of L-type, voltage-dependent Ca2+ channels mediated in large part by PKC.

Original languageEnglish (US)
Pages (from-to)G397-G405
JournalAmerican Journal of Physiology - Gastrointestinal and Liver Physiology
Volume274
Issue number2 37-2
DOIs
StatePublished - 1998

Keywords

  • Bethanechol
  • Diacylglycerol
  • L-type calcium channels
  • Pancreatic β-cells
  • Protein kinase C

ASJC Scopus subject areas

  • Physiology
  • Hepatology
  • Gastroenterology
  • Physiology (medical)

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