Large-Scale Calcium Imaging of Neuronal Activity during Prepulse Inhibition
(A) Top: horizontal projection of gsx1-Gal4 expression (orange) with Mauthner cells (y264-Gal4, green, arrows) that drive fast startle and a pan-neuronal counter-label (HuC, blue), registered to the Zebrafish Brain Browser (ZBB). Yellow box: area imaged in (D). Bottom: coronal cross-section in hindbrain (dotted line in top) shows Gsx1 cell distribution in relation to Mauthner cells. Rostral (R), dorsal (D).
(B) Schematic of large-scale calcium imaging during behavioral prepulse inhibition.
(C) Startle responsiveness (short-latency tail-flip responses, SLCs) during acoustic tests. NS, no stimulus; Pre, prepulse-alone trial. n = 18 larvae. Error bars are SEM. WSR test, ∗p = 0.0002 for suppression of responses on PPI trials compared to pulse-alone trials, confirming that behavioral prepulse inhibition was robustly elicited during imaging.
(D) 2-photon image of nls-GCaMP6s expression in Gsx1 cells (gray, left inset) and mean fluorescence change for 12 prepulse inhibition trials (ΔF, color scale, right inset) for two representative neurons. Activity traces are the mean calcium response to pulse-alone (black, n = 9), prepulse-alone (blue, n = 13), prepulse inhibition (red, n = 12), or no stimulus (orange, n = 11) trials. Shading is SEM. Arrows indicate times of prepulse (small) or pulse (large) stimuli.
(E) Raster plot of normalized GCaMP6s fluorescence change (ΔF/F, color scale), for 655 neurons in a representative experiment from a single larva.
(F) Mean normalized GCaMP6s fluorescence change (ΔF/F, color scale) for all segmented neurons after co-registration, for 4 indicated trial types.
See also Figure S1.