Skeletal muscle is vital for the completion of activities of daily living and is becoming increasingly recognised for its essential roles in whole body metabolism, acting as the largest reservoir for amino acids in the body and a storage point for glucose and lipids. Maintenance of muscle mass is key to the promotion of health, with muscle atrophy as a result of inactivity, disease or ageing, resulting in increased frailty, metabolic comorbidities and mortality. As such, muscle hypertrophy is desirable in the general population due to the rising interest in muscle health and wellbeing, in addition to athletes looking to optimise performance. There have been considerable research efforts aimed at unravelling the regulation of muscle mass, however, many aspects of hypertrophy remain unclear and debated. Understanding the mechanisms and most effective strategies to induce skeletal muscle hypertrophy would offer great resource in combating muscle wasting and add clarity to optimal exercise practices. The use of stable isotope tracers enables the accurate quantification of dynamic metabolism and are key to understanding the mechanisms regulating muscle adaptation. Advances in the application of deuterium oxide (D2O) techniques now permit the simultaneous dynamic measurement of a range of substrates (i.e., protein, lipid, and nucleic acids, along with the potential for OMICs methodologies) with minimal invasiveness further creating opportunities for long-term ‘free living’ measures. As successful muscle adaptation requires a coordinated response of protein turnover, ribosomal biogenesis and satellite cell activity, D2O can provide a more dynamic holistic picture of muscle adaptation.