Functional identification and monitoring of individual α and β cells in cultured mouse islets of Langerhans

Abstract

The aim of the present study was to evaluate, by use of fluorescence microscopy and immunofluorescence stainings, the use of a fluorescent membrane potential sensitive probe as a means to identify and monitor changes in membrane potential of individual cell types in whole islets of Langerhans over time. Our work supports the use of the fluorescent probe bis-(1,3 dibutylbarbituric acid) trimethine oxonol (diBAC₄(3)), in identification of single α and β cells in the periphery of mouse pancreatic islets cultured on extracellular matrix. At a low extracellular glucose concentration (3 mM), heterogeneous staining of the islets was observed. Approximately 97% of the peripheral cells that stained brightly with diBAC₄(3) were glucagon positive. Additional diBAC₄(3) studies, demonstrated that an increase in glucose concentration from 3 to 10 mM is paralleled by repolarization of α cells and depolarization of β cells. This suggests that reciprocity of glucagon and insulin release upon glucose stimulation is coupled to divergent changes in membrane potential of these cell types and supports the use of diBAC₄(3) as a means to detect changes in secretion in both cell types.