Fig. 1

Chronic two-photon calcium imaging of neuronal activity in the IC shell L1 of awake, behaving mice. a Sagittal view showing the cranial window and the in vivo imaging diagram. For the detailed experimental procedures, see the text. Inset: Coronal view showing the central IC (ICC), ICD, ICX, and periaqueductal gray (PAG). b Horizontal view showing the anatomical location of dorsal IC (ICD) and lateral IC (ICX) and the neighboring superior colliculus (SC) and cerebellum. Notice that the IC could be identified by the deep white color. Scale bar: 150 μm. A anterior, L lateral. c Schematic of the experimental set-up. A mouse is head-fixed but free to run on a treadmill with the optical mouse to register its running speed. d Close-field pure tones and click train sound stimuli used. e Example field-of-view (FOV) in the IC shell L1 of awake Vglut2-Cre mouse. f Identical FOV in the IC shell L1 of 1% isoflurane-anesthetized Vglut2-Cre mouse. Stacked images along time axis (3000 images, 5 × 1 × 20 × 30, fps × ITI × stimuli × repeats) were used to generate the standard deviation (SD) figure. More active neurons will have larger SD and brighter color in the figure. 30 μm depth below the dura. Scale bar: 25 μm. g Example fluorescent traces of two excitatory neurons during wakefulness (red) and anesthesia (blue) when stimulated with 70 dB pure tones. h Corresponding spectral tuning curves during wakefulness and isoflurane anesthesia