Specificity of ChR2-YFP expression in DRN 5-HT neurons and efficiency of photostimulation <i>in vitro</i>.

<p><b><i>A</i></b>, Schematics of viral injections. <b><i>B</i></b>, Fluorescence picture of a parasagittal slice showing restricted ChR2-YFP expression in the DRN. <b><i>C</i></b>, Superimposed fluorescence and brightfield images of a ChR2-YFP-expressing coronal slice. Bar: 500 µm. <b><i>D_1–3</i></b>, Confocal pictures of the area delimited by a black rectangle in <i>C</i>. Aq: aquaduct. Bar: 100 µm. <b><i>E_1–3</i></b>, Magnified view of the area delimited by a white rectangle in <i>D</i><b><i>₃</i></b>. Bar: 50 µm. <b><i>F</i></b>, Schematics of patch-clamp recordings in DRN slices. <b><i>G</i></b>, Left: example of a loose cell-attached recording illustrating the protocol used to assess photostimulation thresholds (PTs). Right: blow-up of the trace showing a single photoevoked spike. <b><i>H</i></b>, Spike probability versus incident irradiance for 21 cells. <b><i>I</i></b>, Kernel density estimate of the distribution of logarithmically-scaled PTs (black) and superimposed normalized cumulative sum (red). <b><i>J</i></b>, Spike count per pulse versus pulse duration for 17 cells at twice their PT. Inset: representative firing profiles of 3 different cells identified in the graph by colored arrowheads. <b><i>K</i></b>, Response of a ChR2-YFP cell to trains of repeated light pulses (6 ms) at various frequencies (irradiance set at twice its PT). <b><i>L</i></b>, Average firing rate (± SEM, shaded area) during a 5 s train versus photostimulation frequency at twice the PT (dashed line; n = 29, 28, 25 and 18 cells for 1–2, 5–10, 20 and 50 Hz respectively) and at a higher irradiance (∼5 mW·mm⁻²; solid line, n = 14 and 13 cells for 1–20 and 50 Hz respectively).