Figure 7.

Synaptic NMDAR activity and ${\text{Ca}}^{2+}$ influx drive a reversible dendritic F-actin reorganization from a ring to fiber pattern. (a) Mean distributions of dendritic F-actin rings and fibers using bootstrapping for synaptic stimulation (0${\text{Mg}}^{2+}$/Gly/Bic for 10 min) without (green) or with TTX (black) or APV (red) compared with the low activity high ${\text{Mg}}^{2+}$/low ${\text{Ca}}^{2+}$ condition (blue). (b) Density distribution of the raw data. TTX (1 $\mathrm{\mu }$M) partially but significantly blocks the F-actin remodeling caused by 0${\text{Mg}}^{2+}$/Gly/Bic stimulation, while APV (25 $\mathrm{\mu }$M) blocks it further ($p=1.4\times {10}^{-3}$ and $p=2.5\times {10}^{-5}$, respectively). (c) Representative images of neurons treated with 0${\text{Mg}}^{2+}$/Gly/Bic with (left) and without (right) APV segmented with our deep learning based approach. (d) Mean distributions of dendritic F-actin rings and fibers using bootstrapping for 2 min high ${\text{K}}^{+}$ stimulation (1.2 mM ${\text{Ca}}^{2+}$, orange) or with 2.4 mM ${\text{Ca}}^{2+}$ (violet) or with 1.2 mM ${\text{Ca}}^{2+}$ and 50 $\mathrm{\mu }$M ${\text{Cd}}^{2+}$ (cyan). The red circles indicate the same high ${\text{K}}^{+}$ stimulation (1.2 mM ${\text{Ca}}^{2+}$) condition followed by 15 min wash in high ${\text{Mg}}^{2+}$/low ${\text{Ca}}^{2+}$. The F-actin remodeling is ${\text{Ca}}^{2+}$-dependent and reversible, at least partially within 15 min. (e) Density distribution of the raw data. (f) Representative images of neurons treated with high ${\text{K}}^{+}$ stimuli with (left) and without (right) ${\text{Cd}}^{2+}$ segmented with our deep learning based approach. Statistical analysis performed with a randomization test (see Materials and Methods). Number of independent cultures (N) and number of neurons (n): high ${\text{Mg}}^{2+}$/low ${\text{Ca}}^2+$ $N=9$, $n=86$, 0${\text{Mg}}^{2+}$/Gly/Bic $N=8$, $n=70$; 0${\text{Mg}}^{2+}$/Gly/Bic $1\mathrm{\mu }$M TTX $N=2$, $n=20$; 0${\text{Mg}}^{2+}$/Gly/Bic + $25\mathrm{\mu }$M APV $N=2$, $n=20$; high ${\text{Mg}}^{2+}$/low ${\text{Ca}}^{2+}$ $N=9$; high ${\text{K}}^{+}/1.2$ mM ${\text{Ca}}^{2+}$ $N=9$, $n=83$; high ${\text{K}}^{+}/2.4$ mM ${\text{Ca}}^{2+}$ $N=2$, $n=20$; high ${\text{K}}^{+}/50\mathrm{\mu }$M ${\text{Cd}}^{2+}$ $N=2$, $n=22$; high ${\text{K}}^{+}/15$ min wash high ${\text{Mg}}^{2+}$ $N=3$, $n=32$. For the raw images without overlay see Supplementary Fig. 14.