Nitrogen (N) niche differentiations play an important role in community structure and biogeochemical cycling in terrestrial ecosystems. However, very few studies have examined how plant N strategies specialize via trade-offs between N acquisition and use abilities under natural field conditions. A field experiment was conducted to investigate N strategies by coexisting grass species using the in-situ stable isotope labeling technique. We injected 15N–labeled nitrate, 15N–labeled ammonium, and 13C-15N-labeled glycine solutions in early and late vegetative growing seasons. Shoot δ15N and N concentration were measured to determine the N uptake and use abilities of 4 common species. All plant species preferred to take up nitrate (the dominant N form) over ammonium and glycine, and N-acquisition capacity varied with temporal variation of soil N. However, the dominant species was more N-conservative than less-dominant species and had lower overall N uptake rates (shoot 15N excess 48 hours after injection) and higher N use efficiencies (aboveground biomass : N ratio). The different N strategies may reflect mixed effects of environmental filtering and interspecific competition and have significant implications for species coexistence as well as for ecosystem functions such as nutrient cycling.