Acute perturbation of mTORC1 affects muscle fiber size. Soleus muscle was electroporated with plasmids encoding shRNA directed to transcripts encoding CD4 (Cd4), raptor (Rptor) or TSC2 (Tsc2). Plasmids encoding NLS-GFP were co-electroporated to label transfected fibers. After four to six weeks, muscle fiber size was determined by staining mid-belly cross-sections with Alexa-594-labeled wheat germ agglutinin (red). Transfected muscle fibers were identified by the expression of nuclear-localized GFP (green; white asterisks). The experimental paradigms used were innervated muscle (A, B), reinnervated muscle after nerve crush (C, D) and denervated muscle (E, F). Quantifications (B, D, F) of cross-sectional area (CSA) of muscle fibers in each paradigm are given relative to CSA of neighboring, GFP-negative, non-electroporated fibers. Electroporation of plasmids encoding shRNA to Cd4 served as control. Scale bars (A, C, E) = 50 μm. Bars (B, D, F) represent mean ± SEM (N ≥3 mice and N ≥200 fibers were measured in each). In case of innervated muscles treated with shRNA to Tsc2 and with rapamycin (Tsc2 + Rapa) and denervated muscles electroporated with shRNA to Cd4, data represent mean ± SD (N = 2). P-values are ***P <0.001; **P <0.01; *P <0.05. Unless otherwise indicated, significance was determined compared to the control (shRNA to Cd4).