The Tibetan Plateau (TP) is the headwater of the Yangtze, Yellow, and the transboundary Yarlung Tsangpo, Lancang, and Nujiang Rivers, providing essential and pristine freshwater to around 1.6 billion people in Southeast and South Asia. However, the temperature rise TP has experienced is almost three times that of the global warming rate. The rising temperature has resulted in glacier retreat, snow cover reduction, permafrost layer thawing, and so forth. Here we show, based on the longest observed streamflow data available for the region so far, that changing climatic conditions in the TP already had significant impacts on the streamflow in the headwater basins in the area. Our analysis indicated that the annual average temperature in the headwater basins of these five major rivers has been rising on a trend averaging 0.37 °C·decade⁻¹ since 1998, almost triple the rate before 1998, and the change of streamflow has been predominantly impacted by precipitation in these headwater basins. As a result, streamflow in the Yangtze, Yarlung Tsangpo, Lancang, and Nujiang River headwater areas is on a decreasing trend with a reduction of flow ranging from 3.0–5.9 × 109 m³·decade⁻¹ (−9.12% to −16.89% per decade) since 1998. The increased precipitation in the Tangnahai (TNH) and Lanzhou (LZ) Basins contributed to the increase of their streamflows at 8.04% and 14.29% per decade, respectively. Although the increased streamflow in the headwater basins of the Yellow River may ease some of the water resources concerns, the decreasing trend of streamflow in the headwater areas of the southeastern TP region since 1998 could lead to a water crisis in transboundary river basins for billions of people in Southeast and South Asia.