Calcium modulation of exocytosis-linked plasma membrane potential oscillations in INS-1 832/13 cells

Abstract

In the presence of high glucose or pyruvate, INS-1 832/13 insulinoma cells undergo stochastic oscillations in plasma membrane potential (Δψp) leading to associated fluctuations in cytosolic free Ca2+ concentration ([Ca2+]c). Oscillations are not driven by upstream metabolic fluctuations, but rather by autonomous ionic mechanisms, the details of which are unclear. We have investigated the nature of the oscillator, with simultaneous fluorescence monitoring of Δψp, [Ca2+]c and exocytosis at single-cell resolution, combined with analysis of the occurrence, frequency and amplitude of Δψp oscillations. Oscillations were closely coupled to exocytosis, indicated by coincident synaptopHluorin fluorescence enhancement. L-type Ca2+ channel inhibitors enhanced Δψp and [Ca2+]c oscillation frequency in the presence of pyruvate, but abolished the sustained [Ca2+]c response following KCl depolarization. The L-type Ca2+ channel inhibitor isradipine did not inhibit oscillation-linked exocytosis. The T-type Ca2+ channel inhibitor NNC 55-0396 inhibited Δψp and [Ca2+]c oscillations, implying that T-type Ca2+ channels trigger oscillations and consequent exocytosis. Since distinct ion channels operate in oscillating and non-oscillating cells, quantitative analysis of Δψp and [Ca2+]c oscillations in a β-cell population may help to improve our understanding of the link between metabolism and insulin secretion.