Variations to the global wetland CH₄ source strength in response to changes in orbital insolation patterns and atmospheric CO₂ concentration ([CO₂]a) are hypothesized to play an important role in determining glacial-interglacial variations in atmospheric CH₄ concentration ([CH₄]a). Here the interactive effects of temperature, a major controlling variable determining wetland CH₄ flux, and the low [CO₂]a of glacial intervals are investigated for the first time. We measured the temperature dependence of CH₄ emissions from replicated mesocosms (n = 8 per CO₂ treatment) collected from a minerotrophic fen and an ombrotrophic bog incubated in either ambient (c. 400 ppm) or glacial (c. 200 ppm) [CO₂]a located in the United Kingdom. CH₄ fluxes were measured at 5°C, 10°C, 15°C, 20°C, and 25°C and then in reverse order over a 20 day period under each [CO₂]a treatment. Results showed that the Q10 temperature response of CH₄ emissions from the Carex/Juncus-dominated fen declined significantly by approximately 39% under glacial [CO₂]a (ambient [CO₂]a = 2.60, glacial [CO₂]a = 1.60; P 0.05). This contrasting response may be linked to differences in plant species assemblage and the varying impact of CO₂ starvation on plant productivity and carbon availability in the rhizosphere. Furthermore, our results provide empirical evidence to support recent model-based indications that glacial-interglacial variations in [CH₄]a may be explained by changes in wetland CH₄ source strength in response to orbitally forced changes in climate and [CO₂]a.