Single cell and subcellular measurements of intracellular Ca2+ concentration

McCarron, John G. and Olson, Marnie L. and Chalmers, Susan and Girkin, John M.; Lambert, David G. and Rainbow, Richard D., eds. (2013) Single cell and subcellular measurements of intracellular Ca2+ concentration. In: Calcium Signaling Protocols. Methods in Molecular Biology, 937 . Humana Press, pp. 239-251. ISBN 978-1-62703-086-1

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Increases in bulk average cytoplasmic Ca2+ concentration ([Ca2+]c) are derived from the combined activities of many Ca2+ channels. Near (<100 nm) the mouth of each of these channels the local [Ca2+]c rises and falls more quickly and reaches much greater values than occurs in the bulk cytoplasm. Even during apparently uniform, steady-state [Ca2+] increases large local inhomogeneities exist near channels. These local increases modulate processes that are sensitive to rapid and large changes in [Ca2+] but they cannot easily be visualized with conventional imaging approaches. The [Ca 2+] changes near channels can be examined using total internal reflection fluorescence microscopy (TIRF) to excite fluorophores that lie within 100 nm of the plasma membrane. TIRF is particularly powerful when combined with electrophysiology so that ion channel activity can be related simultaneously to the local subplasma membrane and bulk average [Ca2+]c. Together these techniques provide a better understanding of the local modulation and control of Ca2+ signals.