Calcium measurements in organelles with Ca2+-sensitive fluorescent proteins.
The recent improvement in the design and use of genetically encoded fluorescent Ca2+ indicators should foster major progress in three aspects of Ca2+ signaling. At the subcellular level, ratiometric probes with expanded dynamics are now available to measure accurately the local Ca2+ changes occurring within specific cell compartments. These tools will allow to determine precisely the role of organelles and of cellular microdomains in Ca2+ handling. At the cellular level, the permanent labeling offered by the genetic probes enables large-scale, long-term Ca2+ measurements with robotic multiplexing technologies such as fluorescence plate readers or automated microscopes. This opens the way to large-scale pharmacological or genetic screens based on organelle-specific functional assays. At the whole animal level, probes with improved dynamics and reduced interference with endogenous proteins will allow to generate transgenic animals bearing Ca2+ sensitive indicators in specific cells and tissues. With this approach, Ca2+ signals can be recorded in neurons, heart, and pancreas of live animals during physiological and pathological stimulations. In this chapter, I will review the progress made in the design and use of genetic Ca2+ indicators, and discuss how these new tools provide an opportunity to challenge several unsolved questions in Ca2+ signaling.
Demaurex
Nicolas
N
Department of Cell Physiology and Metabolism, University of Geneva, 1 rue Michel-Servet, CH-1211 Geneva 4, Switzerland. nicolas.demaurex@medecine.unige.ch
D000818:Animals; D002118:Calcium / Q000032:analysis* / Q000737:chemistry* / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015388:Organelles / Q000737:chemistry* / Q000235:genetics
D049452:Green Fluorescent Proteins; D002118:Calcium