ABSTRACT
Fiber photometry is a sensitive and easy way to detect changes in fluorescent signals. The combination of fiber photometry with various fluorescent biomarkers has substantially advanced neuroscience research over the last decade. Despite the wide use of fiber photometry in biomedical fields, the lack of a detailed and comprehensive protocol has limited progress and sometimes complicated the interpretation of data. Here, we describe detailed procedures of fiber photometry for the long-term monitoring of neuronal activity in freely-behaving animals, including surgery, apparatus setup, data collection, and analysis.
Subject(s)
Animals , Female , Male , Mice , Brain , Metabolism , Calcium Signaling , Neurons , Metabolism , Neurosurgical Procedures , Optical Fibers , Optical Imaging , Methods , Photometry , MethodsABSTRACT
Objective As the photostability of calcium ions (Ca2+) indicators is an important property for indicating the temporal features of cytosolic Ca2+ in cells, this study aims to quantitatively measure the light-induced fluorescence enhancement in cells stained with Ca2+ indicators. Methods Five cell lines, MC3T3-E1, RAW264.7, MLO-Y4, MEF3T3 and HEK293, were exposed to the light with five levels of optical power, respectively, so as to investigate the light induced responses of two commonly-used Ca2+ indicators, Fluo-4 AM and Oregon green. The light-induced fluorescence enhancement, the succeeding photobleaching and the thapsigargin (TG)-induced responsive peak followed by were observed. The characteristic parameters of responsive peaks were further analyzed. Results Light with higher power level would induce the fluorescence enhancement for both Fluo-4 AM or Oregon green, while the responsive percentage as well as the magnitude and time span of light-induced peak of Oregon green-stained cells were significantly lower than those of Fluo-4 AM-stained cells. Conclusions The use of Oregon green with low power level light shows better photostability to indicate the intracellular Ca2+.