The responsiveness of the cerebrovasculature to alterations in arterial content of carbon dioxide (PCO2) is a common functional test to assess brain health (i.e., CBF-CO2 responsiveness, also referred to as cerebrovascular reactivity (CVR)). Regular exercise has a positive effect on brain function, and a greater CVR is associated with higher aerobic fitness. Further, an attenuated CVR has been observed in disease populations with impaired brain function (e.g., dementia). The most common and simplest test to assess CVR is the inhalation of a predefined concentration of CO2 via a Douglas bag open circuit (e.g., 5% CO2 in air). This test can be performed and measured using functional magnetic resonance imaging (fMRI), transcranial Doppler (TCD) and/or Duplex Doppler. However, the calculated CVR measure is derived differently between approaches. This study aims to compare CVR measures obtained using these three approaches in active/sedentary and young/old groups where differences in CVR are expected. Ten young healthy volunteers (6 active and 4 sedentary) have been recruited so far, with the aim of recruiting 40 (active/sedentary; young/old). Following full screening and a fitness test, participants completed a gas familiarisation visit. The gas challenge consisted of 4-min room air baseline, followed by two 4-min periods of CO2 inhalation (5% and 7% in air) via the open-circuit Douglas bag method with a 4-min recovery period in-between (room air). Participants attended two experimental sessions on separate days (randomised and counter-balanced). During a session, CVR was either measured using fMRI (DABS sequence to obtain simultaneous blood-oxygen level dependent and perfusion responses) or TCD (measures of middle cerebral artery flow velocity) and Duplex Doppler (measures of vessel diameter and velocity in internal carotid artery). Breath-by-breath measures of end-tidal CO2 were obtained in both visits. fMRI and Doppler CVRs were calculated using a linear regression including baseline and both stimulus concentrations and separately for each stimulus concentration (5% and 7% CO2). Preliminary analysis indicates that there are differences between these CVR measures when comparing between active and sedentary groups; however, initial findings also indicate that there are differences between the neuroimaging methodologies used (e.g., CVR calculated from MRI/TCD/Duplex) as well as variations within the methodologies (e.g., 5% versus 7% CO2 and methods of data extraction). Ongoing data collection and analysis will enable us to better understand these methodological differences. More research is needed to investigate how these measures differ and relate to health and disease, how they vary during healthy ageing, and how they change in response to a clinical intervention targeting improved brain health.