Movie S1.

Photorescue of mGlu₅ receptors by 1PE and 2PE in HEK cells. Video shows GCaMP6s fluorescence in cultured HEK cells co-transfected with mGlu5 receptors as in Fig. 3. Cell in the middle corresponds to cell indicated by arrow in Fig. 3B and Ca²⁺ trace in Fig. 3C, and responds with Ca²⁺ oscillations in all conditions. Intracellular events are recorded at conditions indicated by labels (“agonist” = 3 μM quisqualate; ‘+ alloswitch/no light’ = 1 μM alloswitch supplemented in bath in addition to agonist; ‘1PE’ = 405 nm laser diode, 2 μW; ‘2PE’ = NIR pulsed laser, 780 nm, 20 mW) (SI Appendix, Materials and Methods 5.2). Photostimulation periods start on frame 6 after label, and last 60 frames each. GCaMP6s fluorescence is excited by raster scanning with a 488 nm Argon laser (100 nW, 343 ms/frame). Frame rate is 0.2 Hz during imaging alone, 0.17 Hz during 1P photostimulation, and 0.12 Hz during 2P photostimulation. Picture size is 62×62 μm.