Net CO2 assimilation rates and chlorophyll a fluorescence were measured on leaves of an ABA-unresponsive poplar (Populus trichocarpa × koreana cv. Peace) during a period of drought. Stomata of fully expanded leaves of ‘Peace’ partly closed during drought, while those of Populus euramericana cv. Robusta closed almost completely. The measured data were used to calculate total light-driven electron flows and to derive an estimate of a CO2 mole fraction in the chloroplasts. Two major results were obtained. (i) In well watered plants, photosynthesis operated at lower CO2 mole fractions in the chloroplasts of cv. Peace than in those of cv. Robusta, whereas CO2 mole fractions in the substomatal spaces were of the same order of magnitude. We concluded that a higher resistance to CO2 influx in the mesophyll contributed to the lower net assimilation rates in ‘Peace’. (ii) Drought induced an important decrease in CO2 availability in ‘Robusta’ but not in ‘Peace’. This suggested that reduced CO2 influx was a major cause of the limitation of net CO2 assimilation during drought in ‘Robusta’, but not in ‘Peace’, where drought probably reversibly reduced the apparent carboxylation efficiency of Rubisco. Measurements of O2 evolution under saturating CO2 supported this view, as photosynthesis decreased in droughted plants of ‘Peace’ but not of ‘Robusta’. Moreover, estimates of the 13C/12C isotope ratio in the leaves of both cultivars showed drought-related decreases in discrimination, which, in the case of ‘Peace’, could only be explained by changes in activity of the photosynthetic carbon reduction cycle.