The influences of grazing exclusion on plant species composition and intrinsic water-use efficiency (W_i) of alpine grasslands on the Northern Tibet Plateau are not well understood. We conducted a multisite transect field survey across the three main alpine ecosystems (meadow, steppe, and desert steppe) with nine pairs of grazing-excluded and adjacent open grazed pastures. Short-term grazing exclusion (started in 2006) did not result in significant changes in nutrients or bulk density of the surface soils (0–15 cm), but it slightly changed the aboveground biomass (AGB) and coverage at both community and species levels. Community foliar δ¹³C and W_i differed among alpine grassland types, with values for steppe being similar to those for meadow and with desert steppe values being higher than both of these. Foliar δ¹³C and inferred W_i differed among the dominant species and varied negatively with precipitation and positively with temperature in 2010. These results confirm that there is an environmentally selective effect on the replacement of dominant species. There was no evident difference in foliar δ¹³C or W_i between grazing-excluded and open grazed sites, but there was a slight increase of AGB and coverage in grazing-excluded sites compared to open grazed ones at the species and community levels. These results indicated that grazing exclusion may have no clear influence on the physiological processes related to foliar water usage at the species level, but may have a cumulative effect on the carbon–water balance at the community level. Slight changes in linear regressions of foliar δ¹³C and W_i plotted across climatic gradients indicated that grasslands under grazing management might be more sensitive to regional climatic changes.