TY - JOUR
T1 - Imaging large cohorts of single ion channels and their activity
AU - Hiersemenzel, Katia
AU - Brown, Euan R
AU - Duncan, Rory R
PY - 2013
Y1 - 2013
N2 - As calcium is the most important signaling molecule in neurons and secretory cells, amongst many other cell types, it follows that an understanding of calcium channels and their regulation of exocytosis is of vital importance. Calcium imaging using calcium dyes such as Fluo3, or FRET-based dyes that have been used widely has provided invaluable information, which combined with modeling has estimated the subtypes of channels responsible for triggering the exocytotic machinery as well as inferences about the relative distances away from vesicle fusion sites these molecules adopt. Importantly, new super-resolution microscopy techniques, combined with novel Ca(2+) indicators and imaginative imaging approaches can now define directly the nano-scale locations of very large cohorts of single channel molecules in relation to single vesicles. With combinations of these techniques the activity of individual channels can be visualized and quantified using novel Ca(2+) indicators. Fluorescently labeled specific channel toxins can also be used to localize endogenous assembled channel tetramers. Fluorescence lifetime imaging microscopy and other single-photon-resolution spectroscopic approaches offer the possibility to quantify protein-protein interactions between populations of channels and the SNARE protein machinery for the first time. Together with simultaneous electrophysiology, this battery of quantitative imaging techniques has the potential to provide unprecedented detail describing the locations, dynamic behaviors, interactions, and conductance activities of many thousands of channel molecules and vesicles in living cells.
AB - As calcium is the most important signaling molecule in neurons and secretory cells, amongst many other cell types, it follows that an understanding of calcium channels and their regulation of exocytosis is of vital importance. Calcium imaging using calcium dyes such as Fluo3, or FRET-based dyes that have been used widely has provided invaluable information, which combined with modeling has estimated the subtypes of channels responsible for triggering the exocytotic machinery as well as inferences about the relative distances away from vesicle fusion sites these molecules adopt. Importantly, new super-resolution microscopy techniques, combined with novel Ca(2+) indicators and imaginative imaging approaches can now define directly the nano-scale locations of very large cohorts of single channel molecules in relation to single vesicles. With combinations of these techniques the activity of individual channels can be visualized and quantified using novel Ca(2+) indicators. Fluorescently labeled specific channel toxins can also be used to localize endogenous assembled channel tetramers. Fluorescence lifetime imaging microscopy and other single-photon-resolution spectroscopic approaches offer the possibility to quantify protein-protein interactions between populations of channels and the SNARE protein machinery for the first time. Together with simultaneous electrophysiology, this battery of quantitative imaging techniques has the potential to provide unprecedented detail describing the locations, dynamic behaviors, interactions, and conductance activities of many thousands of channel molecules and vesicles in living cells.
U2 - 10.3389/fendo.2013.00114
DO - 10.3389/fendo.2013.00114
M3 - Article
C2 - 24027557
SN - 1664-2392
VL - 4
SP - 114
JO - Frontiers in Endocrinology
JF - Frontiers in Endocrinology
ER -