Figure 2.

Cooperative functions of Ca_V1.1 and β-catenin in regulation of AChR clustering in E14.5 diaphragms

(A) Mouse whole-mount right diaphragm preparations stained with anti-synapsin (red) and α-BTX (green) at E14.5 to label motor nerve branches and AChR clusters, respectively, in control, Ca_V1.1^−/−, HSA-β-cat^−/−, and HSA-β-cat^−/−; Ca_V1.1^−/− mice. The borders of the AChR cluster band are indicated with white dashed lines.

(B) Framed areas in (A) demonstrating the stepwise deterioration of AChR clustering in control, single-, and double-knockout diaphragms, respectively, captured with a higher magnification objective.

(C) Further 4× magnification showing α-BTX staining. HSA-β-cat^−/−; Ca_V1.1^−/− mice display severely disrupted AChR clustering. Arrows show some of the AChR clusters with a beads-on-a-string-like morphology in HSA-β-cat^−/− mice in (B and C). Scale bars, 200 μm for (A), 100 μm for (B), and 20 μm for (C).

(D–F) Quantification of AChR cluster size (D), AChR coverage in muscle fibers at the endplate region (E), and muscle fiber thickness (F) in control, Ca_V1.1^−/−, HSA-β-cat ^−/−, and HSA-β-cat^−/−; Ca_V1.1^−/− diaphragms. N ≥ 5 diaphragms from ≥5 litters; mean ± SEM; one way ANOVA: F_(3,34) = 15.11; p < 0.0001 for (D); ANOVA: F_(3,42) = 23.80; p < 0.0001 for (E); ANOVA: F_(3,20) = 3.021; p = 0.0538 for (F). Tukey's multiple comparison test: ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ^n.sp > 0.05.