Next-generation GRAB sensors for monitoring dopaminergic activity in vivo

Abstract

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 .

Genetically encoded sensors enable micro- and nano-scopic decoding of transmission in healthy and diseased brains

“…Two families of GPCR-based dopamine (3,4-dihydroxyphenethylamine or DA) sensors, human D 2 dopamine receptor-based GRAB DA [ 25 , 36 ] and human D 1 dopamine receptor-based dLight [ 26 , 37 ] sensors, are engineered (Table 1 ). GRAB DA1m , GRAB DA1h , GRAB DA2m , and GRAB DA2h are the green versions of genetically encoded DA sensors, while rGRAB DA1m and rGRAB DA1h belong to the first red versions of genetically encoded neuromodulatory transmitter sensors [ 25 , 36 ]. dLight sensors also have multiple versions with somewhat different properties [ 26 , 37 ].…”

“…dLight sensors also have multiple versions with somewhat different properties [ 26 , 37 ]. The top-performing GRAB DA2m and dLight 1 registered robust dopaminergic signals from populations of cells in the striatum [ 32 , 36 ] and basal amygdala [ 38 ], which receive the heaviest dopaminergic innervations with probably largest dopaminergic signals [ 38 – 40 ]. A recent addition is a D 2 dopamine receptor-based red version of genetically encoded DA sensor, RdLight 1 [ 41 ], further expanding the toolbox for DA sensors.…”

“…A recent addition is a D 2 dopamine receptor-based red version of genetically encoded DA sensor, RdLight 1 [ 41 ], further expanding the toolbox for DA sensors. Direct experimental comparison showed that GRAB DA2m produced slightly larger single-cell dopaminergic responses with a better signal-to-noise ratio compared to dLight 1 in the same striatal preparation [ 32 , 36 ]. While the extremely low baseline dLight 1 fluorescence often precludes identification of individual expressing cells, dLight 1 appears to have slightly higher selectivity over NE than GRAB DA2m [ 26 , 36 ].…”

“…Direct experimental comparison showed that GRAB DA2m produced slightly larger single-cell dopaminergic responses with a better signal-to-noise ratio compared to dLight 1 in the same striatal preparation [ 32 , 36 ]. While the extremely low baseline dLight 1 fluorescence often precludes identification of individual expressing cells, dLight 1 appears to have slightly higher selectivity over NE than GRAB DA2m [ 26 , 36 ]. It remains unclear whether GRAB DA2m and dLight 1 are capable of detecting modest single-cell dopaminergic signals at other brain areas, such as the prefrontal cortex and hippocampus.…”

Genetically Encoded Biosensors Based on Fluorescent Proteins

“…For example, cp-based sensors detecting the activity of the receptors for acetylcholine [ 106 ], norepinephrine [ 107 ], adenosine [ 108 ], and serotonin [ 109 ] were developed, allowing the monitoring of spatiotemporal activity of various neurotransmitter receptors. In addition to green sensors, red dopamine receptor sensors have recently been developed utilizing cp-mApple [ 110 , 111 ], and thereby different neurotransmitter signals can be simultaneously monitored in living neurons and animals.…”