Extended Data Fig. 1: Neuronal hyperactivity and disruption of calcium homeostasis in CMA-deficient neurons.
a, Extracellular recordings of burst percentage (left) and network burst percentage (right) of L2AKO and control (Ctrl) primary neurons (obtained from female and male pooled brains) in MEA plates at DIV10. Open and closed circles indicate wells from the two independent experiments. b–g, Quantification of number of spikes, number of bursts, average burst frequency (b), average number of spikes per burst (c), burst percentage (d), network burst frequency, number of spikes per network burst, network burst percentage (e), mean firing rate (f) and synchrony index (g) of L2AKO and Ctrl primary neurons in MEA plates at DIV15. Open and closed circles indicate wells from the two independent experiments. h, Ratiometric measurement of cytosolic Ca2+ dyes, Fluo-4-AM (F1) and Fura-Red (F0), to quantify intracellular Ca2+-release following addition of ionomycin (1.5 μM) at indicated times marked by the vertical dotted lines. Imaging performed in Ca2+-free solution in DIV12 L2AKO and Ctrl primary neurons (female and male pooled). F1/ F0 ratiometric quantification (left) and area under the curve (AUC) (right) are shown Boxes: median and 25th and 75th percentiles; whisker ends: 25th and 75th percentiles ±1.5 times the IQR. i,j, Cytosolic Ca2+ quantified in female (i) and male (j) mouse primary neurons using Fluo-4-AM following KCl-induced depolarization F1quantification (left) and AUC (right) are shown. Boxes: median and 25th and 75th percentiles; whisker ends: 25th and 75th percentiles ±1.5 times the IQR. k, Baseline Ca2+ quantified in female and male primary neurons using Fluo-4-AM. From left to right in each graph: n = [12] (a–g) wells from 2 independent experiments per group; n = [3] (h), [6] (i), [5 Ctrl, 7 L2AKO] (j) independent experiments per group; n = [10, 8] (k left), [8, 11] (k right) wells from 3-4 independent experiments per group. Two-tailed unpaired Student’s t-test (a–g,h right, i right, j right, k), or two-way ANOVA test (h left, i left, j left) are used. Data are presented as mean values ±SEM.
