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

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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.

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