Development of the new crop cultivars with high yield under low nitrogen(N) input is a fundamental approach to enhance agricultural sustainability,which is dependent on the exploitation of the elite germplasm.In the present study,four barley genotypes(two Tibetan wild and two cultivated),differing in N use efficiency(NUE),were characterized for their physiological and biochemical responses to different N levels.Higher N levels significantly increased the contents of other essential nutrients(P,K,Ca,Fe,Cu and Mn),and the increase was more obvious for the N-efficient genotypes(ZD9 and XZ149).The observation of ultrastructure showed that chloroplast structure was severely damaged under low nitrogen,and the two high N efficient genotypes were relatively less affected.The activities of the five N metabolism related enzymes,i.e.,nitrate reductase(NR),glutamine synthetase(GS),nitrite reductase(NiR),glutamate synthase(GOGAT) and glutamate dehydrogenase(GDH) all showed the substantial increase with the increased N level in the culture medium.However the increased extent differed among the four genotypes,with the two N efficient genotypes showing more increase in comparison with the other two genotypes with relative N inefficiency(HXRL and XZ56).The current findings showed that a huge difference exists in low N tolerance among barley genotypes,and improvement of some physiological traits(such as enzymes) could be helpful for increasing N utilization efficiency. Development of the new crop cultivars with high yield under low nitrogen (N) input is a fundamental approach to enhance agricultural sustainability, which is dependent on the exploitation of the elite germplasm. The present study, four barley genotypes (two Tibetan wild and two cultivated, differing in N use efficiency (NUE), were characterized for their physiological and biochemical responses to different N levels .Higher N levels significantly increased the contents of other essential nutrients (P, K, Ca, Fe, Cu and Mn) and the increase was more obvious for the N-efficient genotypes (ZD9 and XZ149). The observation of ultrastructure showed the chloroplast structure was severely damaged under low nitrogen, and the two high N efficient genotypes were relatively less affected. The activities of the five N metabolism related enzymes, ie, nitrate reductase (NR), glutamine synthetase (GS), nitrite reductase (NiR), glutamate synthase (GOGAT) and glutamate dehydrogenase (GDH) all showed the substantial increase with the increased N level in the culture medium. Despite the increased extent differed among the four genotypes, with the two N efficient genotypes showing more increase in comparison with the other two genotypes with relative N inefficiency (HXRL and XZ56). that a huge difference exists in low N tolerance among barley genotypes, and improvement of some physiological traits (such as enzymes) could be helpful for increasing N utilization efficiency.