Conversion of forests to agricultural land or pastures is occurring at a rapid rate in many tropical regions. Amphibians may be particularly susceptible to changes in landscape composition and connectivity because of their physiological characteristics and complex life cycles. We experimentally assessed landscape resistance for the dart-poison frog Oophaga pumilio associated with two prevalent land-cover types, secondary forests and pastures, in the northeastern lowlands of Costa Rica. We measured recapture rates of individuals displaced into forests and into pastures, the effects of microclimate on the movement performance of individuals, and the influence of land-cover type and displacement distance on orientation ability of O. pumilio. Results showed a significant interaction between displacement distance and land-cover type indicating greater resistance to movement experienced by individuals displaced into pastures compared with frogs displaced into forests. Microclimatic conditions in pastures had a detrimental effect on the movement performance of O. pumilio and initial orientation was both distance and habitat dependent. Understanding the magnitude of resistance presented by different land-uses to amphibian dispersal is important for the development of successful conservation strategies in human-altered landscapes.