Figure 5. Primary cilia-driven Sstr3 signaling regulates interneuron morphology and connectivity.

(A–C) Striatal tdTom⁺ INs in Nkx2.1Cre; Arl13b^lox/+; Ai9 (A), Nkx2.1Cre; Arl13b^lox/lox; Ai9 (B), and Nkx2.1Cre; Arl13b^lox/lox; Ai9; Sstr3-GFP (C) brains [P30]. (A′–C′) Higher-magnification images of tdTom⁺ striatal INs from outlined area in panels A–C. (D) Induced Sstr3-GFP expression in primary cilia of Nkx2.1Cre; Arl13b^lox/lox; Sstr3-GFP; Ai9 INs. (E) Altered Sstr3 localization in Arl13b deficient IN primary cilia and re-expression after induction of Sstr3-GFP. Co-labeling of primary cilia with anti-ACIII and anti-Sstr3 antibodies in Nkx2.1Cre; Arl13b^lox/+; Ai9, Nkx2.1Cre; Arl13b^lox/lox; Ai9 and Nkx2.1Cre; Arl13b^lox/lox; Ai9⁺; Sstr3-GFP INs. (F–K) Perisomatic boutons (arrowheads) labeled with tdTom (F–H) and anti-VGAT antibodies (I–K) in Nkx2.1Cre; Arl13b^lox/+; Ai9 (F, I), Nkx2.1Cre; Arl13b^lox/lox; Ai9 (G, J) and Nkx2.1Cre; Arl13b^lox/lox; Sstr3-GFP; Ai9 (H, K) brains [P30]. (L–M) Quantification of tdTom⁺ perisomatic bouton density (L) and size (M). Data shown are mean ± SEM. ^*P<0.05 (One-way ANOVA: F_{2,42 [L]} = 23.0; p = 5.53616E-07; post-hoc p_{[L, lox/+ vs. lox/lox]} = 6.08809E-08, post-hoc p_{[L, lox/lox vs. lox/lox-Sstr3]} = 3.26839E-05; F_{2,42 [M]} = 6.54; p = 0.004; post-hoc p_{[M, lox/+ vs. lox/lox]} = 3.64284E-05, post-hoc p_{[M, lox/lox vs. lox/lox-Sstr3]} = 0.008). n = 15 cells from 3 different brains per group. (N–O) Quantification of VGAT⁺ perisomatic bouton density (N) and size (O). Data shown are mean ± SEM. ^*P<0.05 (One-way ANOVA: F_{2,42 [N]} = 13.35; p = 5.6345E-05; post-hoc p_{[N, lox/+ vs. lox/lox]} = 5.21332E-06, post-hoc p_{[N, lox/lox vs. lox/lox-Sstr3]} = 0.004; F_{2,42 [O]} = 17.68; p = 6.04031E-06; post-hoc p_{[O, lox/+ vs. lox/lox]} = 4.23172E-06, post-hoc p_{[O, lox/lox vs. lox/lox-Sstr3]} = 0.0009). n = 15 cells from 3 different brains per group. (P–U) Patch-clamp electrophysiological recordings of striatal MSNs from Nkx2.1Cre; Arl13b^lox/+; Ai9 (control, n=6), Nkx2.1Cre; Arl13b^lox/lox; Ai9 (mutant, n=10) and Nkx2.1Cre; Arl13b^lox/lox; Sstr3-GFP; Ai9 (rescue, n=4) brains. (P, S) Representative patch-clamp electrophysiological recordings from MSNs showing mIPSCs (P) and mEPSCs (S). (Q, R) Quantification of mIPSC frequency (Q) and amplitude (R). ^*P<0.05 (One-way ANOVA, mIPSC frequency: F_2,20 = 7.7; p = 0.003; post-hoc p_{[ctrl vs. IN cKO]} < 0.01, post-hoc p_{[Rescue vs. IN cKO]} < 0.05; control, n = 6; IN cKO, n = 11; rescue, n = 6). (T, U) Quantification of mEPSC frequency (T) and amplitude (U). (V) Rescue of overall excitatory/inhibitory ratio in striatal MSNs of Nkx2.1Cre; Arl13b^lox/lox; Sstr3-GFP; Ai9 mice. Data shown are mean ± SEM. ^*P<0.05 (One-way ANOVA: F_2,19 = 4.8; p = 0.021; post-hoc p_{[ctrl vs. IN cKO]} < 0.05, p_{[Rescue vs. IN cKO]} < 0.05; control, n = 6; IN cKO, n = 10; rescue, n = 6). Scale bar, 37.6μm (A–C); 18μm (A′–C′); 6.2μm (D); 1.3μm (E); 5μm (F–K). See also Figure S5.