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Muscarinic Agonists Activate Ca2+ Store-operated and -independent Ionic Currents in Insulin-secreting HIT-T15 Cells and Mouse Pancreatic β-Cells

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Abstract

The neurotransmitter acetylcholine, a muscarinic receptor agonist, augments glucose-induced insulin secretion from pancreatic β-cells by depolarizing the membrane to enhance voltage-gated Ca2+ influx. To clarify the electrical events involved in this process, we measured ionic currents from a clonal β-cell line (HIT-T15) and mouse pancreatic β-cells. In whole-cell recordings, the muscarinic agonist carbachol (CCh) dose-dependently and reversibly activated a voltage-independent, nonselective current (whole-cell conductance 24 pS/pF, reversal potential ~-15 mV). The current, which we refer to as I musc, was blocked by atropine, a muscarinic receptor antagonist, and SKF 96365, a nonspecific ion channel blocker. The magnitude of the current decreased by 52% when extracellular Na+ was removed, but was not affected by changes in extracellular Ca2+, confirming that I musc is a nonselective current. To determine if I musc activates following release of Ca2+ from an intracellular store, we blocked intracellular IP3 receptors with heparin. Carbachol still activated a current in the presence of heparin, demonstrating the presence of a Ca2+ store-independent, muscarinic agonist-activated ionic current in HIT cells. However, the store-independent current was smaller and had a more positive reversal potential (~0 mV) than the current activated by CCh under control conditions. This result indicates that heparin had blocked a component of I musc, which likely activates following release of stored Ca2+. Depleting IP3-sensitive calcium stores with thapsigargin also activated a non-selective, SKF 96365-blockable current in HIT cells. The properties of this putative store-operated current were similar to the component of I musc that was blocked by heparin, being voltage-independent and reversing near −30 mV. We conclude that I musc consists of store-operated and store-independent components, both of which may contribute to the depolarizing action of muscarinic agonists on pancreatic β-cells.

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  1. Department of Anatomy, Physiology & Genetics, Uniformed Services University of the Health Sciences, Bethesda, MD 20814, USA

    D. Mears & C. L. Zimliki

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Mears, D., Zimliki, C. Muscarinic Agonists Activate Ca2+ Store-operated and -independent Ionic Currents in Insulin-secreting HIT-T15 Cells and Mouse Pancreatic β-Cells . J. Membrane Biol. 197, 59–70 (2004). https://doi.org/10.1007/s00232-003-0642-y

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