The Garonne is the largest river in the south-west of France, and its drainage basin stretches between the Pyrénées and the Massif Central mountains. Until now, no water stable isotope study has been performed on the whole Garonne river basin which is composed of different geological substrata, and where the water resources are limited during the dry summer period. This study focuses on the Garonne river and its tributaries from the Pyrénées foothill upstream to its confluence with the Lot River downstream. The aim of the study is to determine the origins of the surface waters using their chemical and stable isotopic compositions (18O, D and 13C), to better understand their circulation within the drainage basin and to assess the anthropogenic influences. The Garonne displays a specific 18O seasonal effect, and keeps its Pyrénean characteristics until its confluence with the Tarn River. The difference in the dissolved inorganic carbon (DIC) comes mainly from the change in lithology between the Pyrénées and the Massif Central mountains. Agriculture activity is only detected in the small tributaries. Copyright © 2009 John Wiley & Sons, Ltd.