PIP2 in pancreatic β-cells regulates voltage-gated calcium channels by a voltage-independent pathway.

Phosphatidylinositol- 4,5-bisphosphate (PIP2) is a membrane phosphoinositide that regulates the activity of many ion channels. Influx of calcium primarily through voltage-gated calcium (Ca-V) channels promotes insulin secretion in pancreatic beta-cells. However, whether Ca-V channels are regulated by PIP2, as is the case for some non-insulin-secreting cells, is unknown. The purpose of this study was to investigate whether Ca-V channels are regulated by PIP2 depletion in pancreatic beta-cells through activation of a muscarinic pathway induced by oxotremorine methiodide (Oxo-M). Ca-V channel currents were recorded by the patchclamp technique. The Ca-V current amplitude was reduced by activation of the muscarinic receptor 1 (M1R) in the absence of kinetic changes. The Oxo-M-induced inhibition exhibited the hallmarks of voltage-independent regulation and did not involve PKC activation. A small fraction of the Oxo-M-induced Ca-V inhibition was diminished by a high concentration of Ca2+ chelator, whereas >= 50% of this inhibition was prevented by diC8-PIP2 dialysis. Localization of PIP2 in the plasma membrane was examined by transfecting INS-1 cells with PH-PLC delta 1, which revealed a close temporal association between PIP2 hydrolysis and Ca-V channel inhibition. Furthermore, the depletion of PIP2 by a voltage-sensitive phosphatase reduced Ca-V currents in a way similar to that observed following M1R activation. These results indicate that activation of the M1R pathway inhibits the Ca-V channel via PIP2 depletion by a Ca2+-dependent mechanism in pancreatic beta-and INS-1 cells and thereby support the hypothesis that membrane phospholipids regulate ion channel activity by interacting with ion channels.