Figure 5.

Reduced IICR Activity Increases the Lateral Diffusion of GABA_AR γ2 Subunits in the Presence of Calcineurin Activity

QD-SPT tracking of GABA_AR γ2 subunits in hippocampal neurons. (A, E, I, M, and Q) Examples show QD trajectories (green), reconstructed from recording sequences (15.2 s for I and 38.4 s for others), overlaid with FM4-64 signals (gray) in order to identify synapses. Scale bars, 5 μm. (B, F, J, N, and R) Quantifications of median diffusion coefficients are shown (median D ± 25%–75% Interquartile Range [IQR]). (C, G, K, O, and S) The mean (±SEM) synaptic dwell time is shown. (D, H, L, P, and T) Size of confinement domains is shown (mean ± SEM).

(A–H) Tracking of QD-bound GABA_AR γ2 subunits in neurons exposed to IP₃R blocker 2APB for 0–30 (A–D) or 60–90 (E–H) min. Note that exploration and diffusion coefficient of GABA_AR increased after 2APB application (A, B, E, and F). In contrast, the mean synaptic dwell time (C and G) and size of confinement domains (D and H) decreased and increased, respectively, only after 60 min, but not before 30 min of drug treatment.

(I–L) Increased diffusion and reduced synaptic confinement in IP₃R1KO neurons (KO) compared with WT, analyzed at 15–18 DIV.

(M–P) The blockade of group I mGluRs also led to an increase in diffusion of GABA_AR.

(Q–T) Neurons were exposed to 2APB in the absence or presence of the calcineurin inhibitor Cyclosporin A (CysA) and FK506. Both CysA and FK506 prevented the 2APB-dependent increase in GABA_AR diffusion.

NS, p > 0.05; ^∗p < 0.05, ^∗∗p < 0.01, ^∗∗∗p < 0.005; Mann-Whitney U-test (B, F, J, N, and R) and Welch’s t-test for others. Numbers of QDs analyzed are shown in Table S1. See also Figures S5 and S6.