Fluorescent biosensors illuminate calcium levels within defined beta-cell endosome subpopulations.

Live cell imaging has revealed that calcium ions (Ca 2+ ) pass in and out of many cellular organelles. However, technical hurdles have limited measurements of Ca 2+ in acidic organelles, such as endosomes. Although evidence hints that endosomes play a role in Ca 2+ signaling, direct measurements within endosomal lumina represent one of the final frontiers in organelle imaging. To measure Ca 2+ in a TiVAMP-positive endosome sub-population, the pH-resistant ratiometric Ca 2+ biosensor GEM-GECO1 and the ratiometric pH biosensor mKeima were used. A positive correlation between acidic endosomal pH and higher Ca 2+ was observed within these Rab5a- and Rab7-positive compartments. Ca 2+ concentration in most endosomes was estimated to be below 2 μM, lower than Ca 2+ levels in several other intracellular stores, indicating that endosomes may take up Ca 2+ during physiological stimulation. Indeed, endosomes accumulated Ca 2+ during glucose-stimulation, a condition where endosomal pH did not change. Our biosensors permitted the first measurements revealing a role for endosomes in cellular Ca 2+ homeostasis during physiological stimulation. [ABSTRACT FROM AUTHOR]