Miniaturized O2 sensors with tip sizes down to 3 µm have long been available to plant scientists, enabling studies of intra-tissue O2 profiles and associated temporal dynamics. A new CO2 microsensor has recently been developed. This significant advancement in microsensor technology has enabled us to ‘unlock the doors to a secret chamber’ inside plant tissues; high-resolution spatial and temporal CO2 measurements are now possible. We present the first examples of application of the novel CO2 microsensor, and its use side-by-side with an O2 microsensor. We show that an ‘anoxic core’ developed in the stele of chickpea roots as external O2 supply from the surrounding liquid medium was lowered. Concurrently, pCO2 in the centre of the stele accumulated from 0.75 kPa when external O2 was at air equilibrium and up to 1 kPa when external O2 was lowered to 3.7 kPa. Radial tissue profiles through the roots showed a steep declining gradient towards the root surface and CO2 produced in the root diffused out to the surrounding medium. Using the two microsensors, we simultaneously measured both uptake of O2 and production of CO2 to establish the respiratory quotient (RQ) for roots. With pO2 at air equilibrium in the external liquid medium, a small ‘anoxic core’ had already developed and the RQ was slightly above 1, with RQ increasing to 5 as external pO2 was lowered to 3.7 kPa. The data demonstrate the utility of using CO2 and O2 microsensors side-by-side for simultaneous studies of tissue O2 and CO2 dynamics, to monitor responses of respiratory activity to environmental conditions. Hence, we predict that like the use of O2 microsensors, which have been used to resolve questions related to plant aeration, deployment of the new CO2 microsensor will greatly benefit studies in plant ecophysiology to further unravel intra-tissue CO2 profiles and temporal dynamics.