Study region: Upper Madeira Basin (975,500 km2) in Southern Amazonia, which is suffering a biophysical transition, involving deforestation and changes in rainfall regime. Study focus: The evolution of the runoff coefficient (Rc: runoff/rainfall) is examined as an indicator of the environmental changes (1982–2017). New hydrological insights for the region: At an annual scale, the Rc at Porto Velho station declines while neither the basin-averaged rainfall nor the runoff change. During the low-water period Rc and runoff diminish while no changes are observed in rainfall. This cannot be explained by increase of evapotranspiration since the basin-averaged actual evapotranspiration decreases. To explain the decrease of Rc, a regional analysis is undertaken. While the characteristic rainfall-runoff time-lag (CT) at Porto Velho basin is estimated to 60 days, CT is higher (65–75 days) in the south and lower (50 days) over the Amazon-Andes transition regions. It is found that 1) the southern basin (south of 14 °S) best explains low-level Porto Velho runoff, 2) in the south, rainfall diminishes and the frequency of dry days increases. Both features explain the diminution of the runoff and the Rc in Porto Velho. Moreover, the increasing dryness in the south compensates for the rainfall and frequency of wet days (>10 mm) increase north of 14 °S and explains the lack of basin-averaged rainfall trends of the upper Madeira basin.