Imaging and elasticity measurements of the sarcolemma of fully differentiated skeletal muscle fibres.

This study aimed to describe the three-dimensional structure and the elastic properties of the sarcolemma of adult, fully differentiated, skeletal muscle fibres combining Atomic Force Microscopy (AFM) and optical microscopy. Single fibres were enzymatically dissociated from Flexor Digitorum Brevis of adult mice and were maintained in culture up to 3 weeks. On the sixth day after dissociation, the upper surface of intact fibres, either alive in solution or fixed and kept in solution or fixed and exposed in air, was analysed with AFM. The most prominent features in AFM images were periodic transversal foldings with an interval that corresponded to the sarcomere length. More detailed analysis of the topography profile showed that the depth in the folding decreased with increasing sarcomere length and that the crests of the foldings corresponded to the Z-lines. Minor periodic structures could be detected in the valleys between the major foldings. AFM images also showed deep depressions on the sarcolemma likely corresponding to openings of T tubules and caveolae. Two-dimensional elasticity maps were obtained using AFM as an indenter and showed that the crests of the transversal foldings correspond to higher stiffness regions. This study provides the first complete three-dimensional topography and mechanical characterization of intact, living skeletal muscle fibres and might form the basis for further investigations aimed to compare healthy and dystrophic muscles.