Spectral fiber-photometry derives hemoglobin-absorption changes for accurate measurement of fluorescent sensors

Fiber-photometry is an emerging technique for recording fluorescent sensor activity in the brain. Here we demonstrate significant hemoglobin-absorption artifacts that may be misinterpreted as sensor activity changes. Because hemoglobin exists in nearly every location in the brain and its concentration varies over time, such artifacts could significantly impede the accuracy of photometry recording results. We present a novel use of spectral photometry technique and propose a computational method to quantify photon absorption effect from an activity-independent fluorescent protein, which can be used to derive oxy- and deoxy-hemoglobin concentration changes and correct target sensor activities across spectra. We further demonstrate the utility of this method for delineating brain regional differences in neurovascular transfer functions and shed light on the interpretation of hemodynamic-based neuroimaging data.