Ice-core records of the concentration of atmospheric CO2 and its stable isotope ratio (δ13Ca) have shown that the global C cycle has not remained in steady-state over the past 11000 yr, implying a possible change in vegetation activity over this period. Here we evaluated the ecophysiological responses of the dwarf willow (Salix herbacea) over the past 9000 yr by measuring the stable carbon isotope composition and stomatal characters of a unique, well dated, high-latitude (68 °N) sub-fossil leaf sequence. After correction for corresponding changes in δ13Ca, a 9000-yr record of variations in the ratio of intercellular (ci) to atmospheric (ca) CO2 concentration was established. Intercellular[ratio ]atmospheric CO2 concentration ratios provide a time-integrated indicator of the set-point of leaf gas exchange, and the historical variations revealed in this record have been interpreted as an impact of environmental changes on leaf gas exchange. The sequence shows a progressive fall in ci/ca 9000–3000 yr BP as well as the climatic effects of the Medieval Warm Period, the Little Ice Age and the post-industrial CO2 rise. Leaf stomatal index (proportion of epidermal cells as stomata), but not stomatal density, was significantly (P<0.01) correlated with Holocene atmospheric CO2 variations. A process-based interpretation of the changes in ci/ca was made using two different coupled photosynthesis-stomatal conductance models. Calculated in this way, S. herbacea photosynthetic rates were relatively stable throughout the Holocene whilst stomatal conductance progressively declined. Both, however, showed the marked effects of the Medieval Warm Period and the Little Ice Age. Overall, the results demonstrate that S. herbacea leaf metabolism, like the global C cycle, has not remained in steady state during the Holocene but has responded to changes in atmospheric CO2 concentration and short-term climatic oscillations.