Fig. 5: 3D functional imaging in awake mouse brains.

a, b Orthogonal MIPs of GCaMP6s-labeled neurons at t = 51.48 s reconstructed by traditional LFM and QLFM with a ×20/0.5NA objective, indicating the reduced background with computational optical sectioning (Supplementary Movie 4). More neurons were revealed by QLFM in the mouse cortex with the native objective plane at a depth of ~250 μm. c Temporal traces of the marked neurons in a and b, demonstrating the quantitative calcium responses in QLFM with improved contrast. d Scatter diagram of SBR of each neuron marked in a and b by different methods. Data points from the same neuron obtained by traditional LFM and QLFM are connected by a red line. P value was calculated by the paired, two-tailed t-test (α = 0.05). e, f Orthogonal MIPs of the standard deviation across 2000 volumes imaged at a center depth of 240 μm and 280 μm in the cortex, respectively, under a ×40/1.0 NA water-immersion objective (Supplementary Movie 5). g Orthogonal MIPs of the standard deviation across 500 volumes for GCaMP6f-labeled L2/3 neurons in awake behaving mice. The video was captured with high-speed axial scanning at a step of 50 μm for three planes under a ×20/0.5 NA objective. h Temporal traces of several marked neurons at different depths. i Scatter diagram of SBR for five selected neurons in g. Data points from the same neuron obtained by traditional LFM and QLFM are connected by a red line. P value was determined via the paired, two-tailed t-test (α = 0.05). Source data are provided as a Source data file. Scale bars, 100 μm (a, b, g) and 50 μm (e, f).