Fig. 7: Bidirectional modulation of neuronal function by FacePalm variants.

a Confocal images of hippocampal neurons transduced with FacePalm-17A at baseline (left) and upon incubation with rapamycin for 4 h (right). PSD95 (green), mcherry marker for FacePalm expression (red), and DAPI (blue). Bottom, enlarged view. Scale bar, 5 µm. b Cumulative histogram compares changes in puncta density of PSD95 with FacePalm-17A at baseline (gray) or with rapamycin. Basal, n = 110 processes from 11 cells; +Rapa, n = 63 processes from seven cells from three independent cultures. **p = 0.0089 by using Kolmogorov-Smirnov test. c, d Activation of FacePalm-17B reduces PSD95 puncta density in cultured hippocampal neurons. Format as in (a, b). Scale bar, 5 µm. Basal, n = 127 processes from 11 cells; +Rapa, n = 103 processes from 12 cells from three independent cultures. ** p = 0.0032 using Kolmogorov-Smirnov test. e, f Activation of FacePalm-17C also reduces PSD95 puncta density. Format as in (a, b). Scale bar, 5 µm. Basal, n = 97 processes from nine cells; +Rapa, n = 99 processes from 11 cells from three independent cultures. **p = 0.0032 by Kolmogorov-Smirnov test. ****p < 0.0001 by Kolmogorov-Smirnov test. g Exemplar traces from current clamp recordings show action potentials evoked from hippocampal neurons transduced with Facepalm-17A in response to 400 pA current injection for 1 s. Black trace, basal; Red trace, with rapamycin. h AP firing rate in response to various current injections. Each dot, mean ± s.e.m. n = 12 cells (basal); n = 17 cells (+rapa) from three independent cultures. Activation of FacePalm-17B results in increased AP firing. Exemplar traces (i), and quantification of AP firing rate (j). Each dot, mean ± s.e.m. n = 8 cells (basal); n = 10 cells (+rapa) from three independent cultures. *p < 0.05 by unpaired two-tailed multiple t-tests (p = 0.0259 at 350 pA, p = 0.0219 at 400 pA, p = 0.0126 at 450pA). k, l Activation of FacePalm-17C results in decreased AP firing. Exemplar traces (k), and quantification of AP firing rate (l). Each dot, mean ± s.e.m. n = 12 cells (basal); n = 9 cells (+rapa) from three independent cultures. *p < 0.05, **p < 0.01 by unpaired two-tailed multiple t-tests (p = 0.0317 at 350pA, p = 0.0035 at 400 pA, p = 0.0203 at 450 pA). m Confocal images show hippocampal neurons transduced with PSD95-targeted FacePalm-it at baseline (left) and upon incubation with rapamycin for 4 h (right). Scale bar, 10 µm. n Cumulative histogram compares changes in the puncta density of PSD95 with FacePalm-it targeting PSD95 at baseline (gray) or with rapamycin (red). Basal, n = 241 processes from 37 cells; +Rapa, n = 186 processes from 27 cells from three independent cultures. ***p = 0.002 by Kolmogorov-Smirnov test. Activation of FacePalm-it targeting PSD95 minimally perturbs AP firing rate. Exemplar traces (o), and quantification of AP firing rate (p). Each dot, mean ± s.e.m. n = 9 cells (basal); n = 11 cells (+rapa) from three independent cultures. Source data are provided as a Source Data file.