Glucose-Stimulated Oscillations in Free Cytosolic ATP Concentration Imaged in Single Islet β-Cells

Abstract

Normal glucose-stimulated insulin secretion is pulsatile, but the molecular mechanisms underlying this pulsatility are poorly understood. Oscillations in the intracellular free [ATP]/[ADP] ratio represent one possible mechanism because they would be expected to cause fluctuations in ATP-sensitive K⁺ channel activity and hence oscillatory Ca²⁺ influx. After imaging recombinant firefly luciferase, expressed via an adenoviral vector in single human or mouse islet β-cells, we report here that cytosolic free ATP concentrations oscillate and that these oscillations are affected by glucose. In human β-cells, oscillations were observed at both 3 and 15 mmol/l glucose, but the oscillations were of a longer wavelength at the higher glucose concentration (167 vs. 66 s). Mouse β-cells displayed oscillations in both cytosolic free [Ca²⁺] and [ATP] only at elevated glucose concentrations, both with a period of 120 s. To explore the causal relationship between [Ca²⁺] and [ATP] oscillations, the regulation of each was further investigated in populations of MIN6 β-cells. Incubation in Ca²⁺-free medium lowered cytosolic [Ca²⁺] but increased [ATP] in MIN6 cells at both 3 and 30 mmol/l glucose. Removal of external Ca²⁺ increased [ATP], possibly by decreasing ATP consumption by endoplasmic reticulum Ca²⁺-ATPases. These results allow a model to be constructed of the β-cell metabolic oscillator that drives nutrient-induced insulin secretion.