Soil nitrogen (N) availability is fundamental for N uptake and biomass accumulation by plants and is affected by irrigation management. Deficit irrigation is an efficient strategy to manage soil moisture for crop production. However, responses of N uptake and soil mineral N to deficit irrigation are not yet clear in alfalfa grasslands. A 2-year trial with alfalfa was carried out using two irrigation systems, flood and subsurface drip irrigations, at three irrigation levels: full irrigation, slight water deficit irrigation (70% of full irrigation), and moderate water deficit irrigation (50% of full irrigation) for the whole growth period. The N uptake (Nuptake) and N use efficiency (NUE) of alfalfa, as well as the soil mineral N, were measured. The Nuptake by plant organs decreased with increasing deficit level of irrigation, but slight deficit irrigation barely reduced Nuptake as compared to full irrigation. Deficit irrigation led to a decrease in dry matter yield, especially in the 2nd year, but slight deficit irrigation reduced it little. In contrast, total N concentration in plant organs barely changed with the deficit level of irrigation, suggesting that biomass growth, rather than change in N concentration, may play a major role in regulating Nuptake. The NUE of alfalfa barely changed with the deficit level of irrigation, although there was a decreasing tendency in the 2nd year. There were significant correlations between Nuptake and NUE of alfalfa with soil ammonium N content, suggesting that this form of N is relatively short-lived and nitrate N may be adequate to meet the growth requirement of alfalfa. In practice, it is recommended that slight deficit irrigation should be set up during alfalfa production in the inland arid area of China.