Dopamine (DA) is a central monoamine neurotransmitter involved in many physiological and pathological processes. A longstanding yet largely unmet goal is to measure DA changes reliably and specifically with high spatiotemporal precision, particularly in animals executing complex behaviors. Here we report the development of genetically-encoded GPCR-Activation-Based-DA (GRABDA) sensors that enable these measurements. In response to extracellular DA, GRABDA sensors exhibit large fluorescence increases (ΔF/F0 ~90%) with subcellular resolution, sub-second kinetics, nanomolar to sub-micromolar affinities, and excellent molecular specificity. GRABDA sensors can resolve a-single-electrical-stimulus evoked DA release in mouse brain slices, and detect endogenous DA release in living flies, fish, and mice. In freely-behaving mice, GRABDA sensors readily report optogenetically elicited nigrostriatal DA release and depict dynamic mesoaccumbens DA signaling during Pavlovian conditioning or during sexual behaviors. Thus, GRABDA sensors enable spatiotemporally precise measurements of DA dynamics in a variety of model organisms while exhibiting complex behaviors.

Dopamine (DA) plays a critical role in the brain, and the ability to directly measure dopaminergic activity is essential for understanding its physiological functions. We therefore developed red fluorescent GPCR-activation–based DA (GRAB DA ) sensors and optimized versions of green fluorescent GRAB DA sensors. In response to extracellular DA, both the red and green GRAB DA sensors exhibit a large increase in fluorescence, with subcellular resolution, subsecond kinetics, and nanomolar to submicromolar affinity. Moreover, the GRAB DA sensors resolve evoked DA release in mouse brain slices, detect evoked compartmental DA release from a single neuron in live flies, and report optogenetically elicited nigrostriatal DA release as well as mesoaccumbens dopaminergic activity during sexual behavior in freely behaving mice. Co-expressing red GRAB DA with either green GRAB DA or the calcium indicator GCaMP6s allows simultaneously tracking neuronal activity and dopaminergic signaling in distinct circuits in vivo .

Highlights d Temporal difference (TD) error is a powerful teaching signal in machine learning d Teleport and speed manipulations are used to characterize dopamine signals in mice d Slowly ramping as well as phasic dopamine responses convey TD errors d Dopamine neurons compute TD error or changes in value on a moment-by-moment basis