Ao Dong, Kaikai He, Barna Dudok, Jordan S Farrell, Wuqiang Guan, Daniel J Liput, Henry L Puhl, Ruyi Cai, Jiali Duan, Eddy Albarran, Jun Ding, David M Lovinger, Bo Li, Ivan Soltesz, Yulong Li. bioRxiv 2020.10.08.329169; doi: Online ahead of print.

Endocannabinoids (eCBs) are retrograde lipid neuromodulators involved in many physiologically important processes. However, their release and dynamics in the brain remain largely unknown, in part due to lack of probes capable of reporting real-time eCBs with sufficient spatiotemporal resolution. Here, we developed a new G protein-coupled receptor Activation Based eCB sensor GRABeCB2.0 using the human CB1 cannabinoid receptor and a circular-permutated EGFP. GRABeCB2.0 exhibited proper cell membrane trafficking, ~seconds kinetics, high specificity and robust fluorescence response to eCBs at physiological concentrations. Using GRABeCB2.0, we detected evoked eCB dynamics in both cultured neurons and acute brain slices. Interestingly, we also observed spontaneous compartmental eCB transients that spread ~11 μm in cultured neurons, suggesting locally-restricted eCB signaling. By expressing GRABeCB2.0 in vivo, we readily observed foot-shock elicited and running triggered eCB transients in mouse amygdala and hippocampus, respectively. Lastly, using GRABeCB2.0 in an epilepsy model, we observed a spreading eCB wave following a calcium wave in mouse hippocampus. In summary, GRABeCB2.0 is a powerful new probe to resolve eCB release and dynamics under both physiological and pathological conditions.