Extended Data Fig. 4: Response statistics of the microablated neurons.

a. Summary schematic indicating the average fraction of neurons with their categorization of responsiveness and different cell types. For a subset of mice in each experimental cohort (5/10, 5/10, 7/9 mice in sound responsive cohort, non-sound responsive cohort, control cohort, respectively), we also labeled inhibitory cortical neurons by co-injecting an AAV vector encoding the mDlx enhancer system (AAV-mDlx-NLS-tagBFP) with the AAV vectors expressing GCaMP6m and H2B::mCherry (see also Fig. 5 and Extended Data Fig. 8). Therefore, we were able to identify the excitatory and inhibitory types of microablated and spared neurons. Small rectangles indicate the fraction of microablated neurons for each experimental cohort. The length of horizontal side refers to the fraction of microablated neurons out of all qualified neurons per FOV. The fraction along the vertical side refers to the ratio of excitatory and inhibitory neurons in the microablated neurons. The categorization of responsiveness is also given for an additional experimental cohort in which inhibitory neurons were microablated (see also Fig. 6). Orange: sound responsive ablation; Blue: non-sound responsive ablation; Pink: inhibitory neuron ablation. The fraction along the horizontal side in the pink rectangle refers to the ratio of significantly responsive and unresponsive neurons in microablated inhibitory neurons per FOV. b. Summary table of number of mice, number of FOV, number of neurons and number of averaged microablated neurons per FOV acquired for each experimental cohort. For sound responsive ablation and non-sound responsive ablation, 41.5 ± 1.75 neurons and 40.3 ± 2.40 neurons were targeted per mouse on average, meaning that 81.9 ± 5.5% and 87.5 ± 2.4% were successfully ablated out of the total number of target neurons respectively, based on the criteria described in Methods. c. Cumulative distribution of number of neurons per FOV across all the 7 imaging time points for each experimental cohort. Neurons were included in this analysis after assessing imaging quality (Methods) and removal of high and low category neurons (Fig. 2c, Methods). Two-sided Kolmogorov-Smirnov test between experimental cohorts with FDR correction: p > 0.36. d. Cumulative distribution of fraction of responsive neurons averaged over the three baseline imaging sessions for each FOV. Two-sided Kolmogorov-Smirnov test for fraction of responsive neurons with FDR correction, all pairwise combination of the experimental cohorts among sound responsive ablation, non-sound responsive ablation, control and inhibitory neuron ablation: p > 0.62. e. Spatial distribution of microablated neurons for each experimental cohort. Top: distribution along depth from dura. Bottom: Distribution of number of microablated neurons per FOV. f. Box plots of number of microablated neurons per mouse across experimental cohorts. One-way ANOVA test, p = 0.63. g. Number of high category neurons per mouse, which means number of microablated neurons for sound responsive ablation and number of corresponding neurons for the other cohorts. One-way ANOVA test, p = 0.65. h. Number of low category neurons per mouse, which means number of microablated neurons for non-sound responsive ablation and number of corresponding neurons for the other cohorts. One-way ANOVA test, p = 0.95. i. Response amplitude of microablated neurons averaged during baseline days (3 and 5). The amplitude of non-sound responsive neurons was defined as the average amplitude across stimuli for each neuron. Two-sided t-test with FDR correction between each two of the three experimental cohorts (Sound responsive ablation vs. Non-responsive ablation: p = 2.68 × 10⁻⁹; Sound responsive ablation vs. Inhibitory neuron ablation: 2.99 × 10⁻⁵; Non-responsive ablation vs. Inhibitory neuron ablation: 3.90 × 10⁻⁹). *** p < 0.001. j. Best response amplitude of high category neurons during baseline days. One-way ANOVA test, p = 0.73. k. Response amplitude of low category neurons during baseline days. The amplitude of non-sound responsive neurons was defined as the average amplitude across stimuli for each neuron. One-way ANOVA test, p = 0.14. l. Number of spared neurons per FOV considered for analysis. One-way ANOVA test, p = 0.86. n = 10 mice for sound responsive ablation, n = 10 mice for non-sound responsive ablation, n = 6 mice for inhibitory neuron ablation for f-l, n = 9 mice for control for g,h,j-l. For box plots in f-l, the middle line, the bottom and top edges of the box indicate median, 25th and 75th percentiles, respectively, and the whiskers are the most extreme data points across mice. See Supplementary Table 2. for detailed statistics.