Agroforestry systems are known to provide ecosystem services which differ in quantity and quality from conventional agricultural practices and could enhance rural landscapes. In this study we compared ecosystem services provision of agroforestry and non-agroforestry landscapes in case study regions from three European biogeographical regions: Mediterranean (montado and dehesa), Continental (orchards and wooded pasture) and Atlantic agroforestry systems (chestnut soutos and hedgerows systems). Seven ecosystem service indicators (two provisioning and five regulating services) were mapped, modelled and assessed. Clear variations in amount and provision of ecosystem services were found between different types of agroforestry systems. Nonetheless regulating ecosystems services were improved in all agroforestry landscapes, with reduced nitrate losses, higher carbon sequestration, reduced soil losses, higher functional biodiversity focussed on pollination and greater habitat diversity reflected in a high proportion of semi-natural habitats. The results for provisioning services were inconsistent. While the annual biomass yield and the groundwater recharge rate tended to be higher in agricultural landscapes without agroforestry systems, the total biomass stock was reduced. These broad relationships were observed within and across the case study regions regardless of the agroforestry type or biogeographical region. Overall our study underlines the positive influence of agroforestry systems on the supply of regulating services and their role to enhance landscape structure.