Imaging with organic indicators and high-speed charge-coupled device cameras in neurons: some applications where these classic techniques have advantages

Abstract

Abstract. Dynamic calcium and voltage imaging is a major tool in modern cellular neuroscience. Since the beginning of their use over 40 years ago, there have been major improvements in indicators, microscopes, imaging systems, and computers. While cutting edge research has trended toward the use of genetically encoded calcium or voltage indicators, two-photon microscopes, and in vivo preparations, it is worth noting that some questions still may be best approached using more classical methodologies and preparations. In this review, we highlight a few examples in neurons where the combination of charge-coupled device (CCD) imaging and classical organic indicators has revealed information that has so far been more informative than results using the more modern systems. These experiments take advantage of the high frame rates, sensitivity, and spatial integration of the best CCD cameras. These cameras can respond to the faster kinetics of organic voltage and calcium indicators, which closely reflect the fast dynamics of the underlying cellular events.

Spatial and temporal aspects of neuronal calcium and sodium signals measured with low-affinity fluorescent indicators

“…Optical recordings of Ca 2+ or Na + concentration changes are ultimately indirect measurements of the fluorescence of an indicator, which is a molecule that binds to the ion and changes its photon absorption and emission. While this molecule can be a genetically encoded protein [41], organic indicators are still widely used since they can reveal information that would not be available otherwise [64]. In all cases, the biophysical challenge consists in extracting, from this indirect measurement, the physiologically relevant information.…”