Effects of tricaine and gradual cooling on sensory and motor brain areas. (A) The duration of drug exposure was the same as in our previous experiments. Stimuli were presented alongside calcium imaging, and ceased between recordings, as shown in the schematic. Tricaine (168 mg/L and 84 mg/L) and gradual cooling treatments were applied for 7 min (Exposure). (B) During imaging of the pretectum, the stimulus consisted of alternating moving (7 s) and stationary (12 s) periods in order to determine the motion sensitivity of identified neurons. (C) For experiments in the hindbrain, the stimulus was constantly rotating (18°/s) and alternated between clockwise and counterclockwise motion every 12 s. (D,E) Motion-sensitivity of neurons detected in larvae exposed to tricaine and gradual cooling. Note that cooling treatment reduced the number of detected motion-sensitive cells and their motion-sensitivity, while the corresponding effects during tricaine treatment were less pronounced or absent. In the upper row (% Max. Cells), the number of detected neurons is expressed as a percentage of the maximum number of neurons detected via pixel-wise regressor correlation in any minute of the recording. The absolute number of identified cells in individual recordings is shown via the diameter of the data points. (F,G) The number of stimulus-associated neurons detected in the hindbrain of larvae exposed to either tricaine or gradual cooling was decreased. The dynamic range of eye positions is shown in the top row. Plots in (D–G) differ in style, because in (D,E) we assessed two parameters (motion sensitivity and number of cells, each represented by the circles) and only one parameter (number of cells) in (F,G). Results were binned per phase into baseline, treated, recovery, recording 2 and recording 3 and analyzed via two-way repeated measures ANOVA with Tukey's HSD test, n = 4–7. *p < 0.05, ***p < 0.001.
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