Focal adhesion kinase (FAK) is a mechano-regulated signalling molecule which controls assembly and attachment of sarcomeres to sarcolemmal sites of focal adhesion in culture (costameres) (Quach & Rando 2006; Wilkinson et al 2008; Durieux 2009). We set out to determine whether adjustments in FAK expression and activation status are associated with load-dependent adaptations in size and costamere content of fully developed human and rat skeletal muscle. Loading to human vastus lateralis muscle was modified in healthy male subjects via muscle unloading by bedrest (n=9; 24 yrs, 180 cm, 77 kg) and overload by resistance training (n=6; 23 yrs; 175 cm; 71 kg). Biopsies were collected from m. vastus lateralis under local anaesthesia (2% lignocaine) before and during the course of the intervention, i.e. 7 and 28 days mid and post bedrest and after the 3rd and 27th resistance exercise session. Protein content of FAK, the costamere components meta- and gamma-vinculin, and FAK activation status (i.e. phosphorylation of Y397; FAK-pY397) were assessed by western blotting. FAK protein was localized with confocal microscopy. Full-length FAK protein was overexpressed via electroporation with a cytomegalovirus-driven expression plasmid in soleus muscle of isoflurane-anaesthetized (3% in O2) male Wistar rats which loading state was altered by hindlimb-unloading and reloading as described (Durieux et al 2008). Controlateral muscle being transfected with empty plasmid served as intra-animal control. Human and animal experiments were carried out with permission of the local ethics committee. FAK protein was mainly expressed at the sarcolemma of muscle fibres. Content of costamere components FAK and gamma-vinculin in m. vastus lateralis was not altered after unloading and overloading despite significant changes in muscle cross section mid (+5%, p<0.05, repeated ANOVA) and post resistance training (+10%) and reduced muscle thickness mid (-7%) and post bedrest (-17%). By contrast, FAK-pY397 content was selectively increased after resistance training (+173%) and correlated with meta-vinculin content (r=0.91) which was transiently reduced mid into resistance training (-28%). Changes in meta-vinculin were correlated with alterations in muscle cross sectional area/muscle thickness mid into resistance training (r=0.79) and bedrest (r=-0.86) and altered FAK-pY397 content and concentration, respectively, after resistance training (r=0.91) and mid into bedrest (-0.82). FAK-overexpression in rat m. soleus increased FAK-pY397 content and elevated the content of meta (+84%) but not the gamma vinculin isoform (p=0.23) in correspondence with load-regulated FAK-pY397 content. The load-dependent control of meta-vinculin expression by FAK activity exposes a novel role of costamere turnover during sarcomere remodelling.