The sustainability of changes in well-being achieved via positive interventions is challenged by findings that happiness levels are constrained by a homeostatic set-point. In this paper, we propose that while generally stable, the neurological and psychophysiological bases of well-being demonstrate plasticity. The neurobiological underpinnings of the hedonic component of well-being are first reviewed, demonstrating the value of both central (such as frontal asymmetry) and peripheral (such as heart rate variability) indices. Convergent evidence that certain well-being interventions are capable of modifying subjective, central and peripheral indices of positive affect or regulation of negative affect is then reported, although there is a clear need for longitudinal research to demonstrate the longevity of changes. It is recommended that a multi-level approach to evaluating positive interventions incorporating subjective psychosocial and neurobiological indices of affective change is adopted by researchers in an attempt to identify interventions most likely to achieve sustained positive outcome. Accumulating evidence through rigorous research that positive interventions can enhance psychosocial and neurophysiological factors can provide a compelling case for more widespread dissemination through public health policy.