Figure 4. Typical mapping experiment using nontargeted ChR2.
(A) In acute slices expressing GCaMP6s (green) and conventional ChR2-mRuby2, cells were identified for photostimulation based on responsiveness to widefield ChR excitation (yellow circles). A cell was patched and dye-filled (triangle) and assessed for postsynaptic currents as surrounding cells were stimulated (2.13 mW/µm2 incident power) and GCaMP fluorescence was simultaneously recorded. Scale bar = 100 µm. (B) Changes in GCaMP fluorescence over the experimental timecourse for each cell identified in (A); each cell was stimulated sequentially as described in Figure 3. Signals are the average of four trials. (C) Calcium (blue) responses for a subset of cells reacting to TF stimulation; four trials are overlaid in light blue, with the average in dark blue. Red lines in the calcium traces indicate the onset of stimulation for each cell. The recorded currents for the four trials (black) in the patched and putatively postsynaptic cell are shown expanded below each calcium trace; the shaded red region indicates the 150 ms stimulation epoch. Several types of events were observed: one cell showed a synaptic event coincident with a low amplitude current from direct stimulation of the dendritic arbor of the patched cell, and one cell showed what might be a synaptic event buried within another direct stimulation current of large amplitude (red arrows). Three other cells with a calcium response to TF stimulation and coincident direct stimulation of the patched cell’s dendritic arbor are also shown, along with three cells with calcium transients but no detectable currents in the patched cell. The cell numbers, spatial locations and full calcium traces corresponding to these events are marked in panels (A) and (B).