Almost 90% of agricultural land in South Tyrol is composed of extensively and intensively used meadows und pastures. Land-use in mountain environments has strong influence on water supply in the lower regions, on soil stability and on hydropower. With the Alpine region not only being influenced by large-scale changes in land-use during the last decades, but also being affected by climate change more than the global mean, it is essential to deeply understand the hydrological mechanisms of Alpine grassland ecosystems. Specifically, I was interested in (1) evaluating the impact of different agricultural management types (intensification and extensification) and changing climate (higher mean temperatures, lower precipitation) on grasslands in Alpine environments by performing a common garden experiment during the vegetative seasons of 2015 and 2016; and (2) investigating the use of water sources by grassland plants in relation to spatial and temporal parameters as well as vegetation type. Both studies were conducted in Stubai Valley in Tyrol, Austria, by using smart-field lysimeters, trace gas analysers and pore water samplers in the field and cryogenic distillation, stable isotope ratio mass spectrometry and cavity ring-down laser spectroscopy in the laboratory. I used stable isotope analysis of carbon (13C/12C) and oxygen (18O/16O) of plant biomass to quantify water use efficiency (WUE), photosynthetic activity and stomatal conductance of different grassland species, while hydrogen (2H/1H) and oxygen isotopic signature of water was used to trace the supply of water from the different possible sources (ground water, rain and stream water) into plants. To verify the usability of pore water samplers with porous tips for extraction of soil water bound for analysis of stable isotopes, I also tested their fractionation effects on sampled soil water. Finally, I investigated the changes of isotopic composition in metabolic compounds in wheat plants during droughts and during exposure to pre-industrial and pre-glacial CO2 concentrations. With this approach of combining well-established ecological techniques and state-of-the-art technology, this thesis contributes to a better understanding of hydrology and the future development of agricultural ecosystems in mountain environments in a changing world.