Fig. 4. In vivo osteocyte morphology is affected by ACh signaling disruption in a sexually dimorphic way.

Endogenously expressed GCaMP6f signal in osteocytes was used to evaluate 2D cellular morphology in vivo. (A) There were no differences in the number of cells per area between the groups. (B) Female Chrna1 cKO osteocytes have a slightly lower cell area when compared to GCaMP controls. For both females (C to F) and males (G to L), Chrna1 cKO cells were more elongated and less round than controls. (A to F) Rapsn cKO did not alter 2D osteocyte morphology in females. In Rapsn cKO males, cells were again more elongated (J to L). These results suggest that ACh signaling disruption produces altered osteocyte shape, which could explain the impacts on in vivo osteocyte mechanotransduction. Two-way analysis of variance (ANOVA) with Tukey’s multiple comparison test was performed with cKO and GCaMP controls to determine significance (*P < 0.05; **P < 0.01, ***P < 0.001, ****P < 0.0001) for each metric. N = 4 to 69 cells per mouse, group sample size (GCaMP: 7 to 9, Chrna1: 9 to 10, Rapsn: 12 to 14).