BACKGROUND: A large quantity of nitrogen (N) fertilizer is used for crop production to achieve high yields at a significant economic and environmental cost. Efforts have been directed to understanding the molecular basis of plant responses to N and identifying N-responsive genes in order to manipulate their expression, thus enabling plants to use N more efficiently. No studies have yet delineated these responses at the transcriptional level when plants are grown under chronic N stress and the understanding of regulatory elements involved in N response is very limited. RESULTS: To further our understanding of the response of plants to varying N levels, a growth system was developed where N was the growth-limiting factor. An Arabidopsis whole genome microarray was used to evaluate global gene expression under different N conditions. Differentially expressed genes under mild or severe chronic N stress were identified. Mild N stress triggered only a small set of genes significantly different at the transcriptional level, which are largely involved in various stress responses. Plant responses were much more pronounced under severe N stress, involving a large number of genes in many different biological processes. Differentially expressed genes were also identified in response to short- and long-term N availability increases. Putative N regulatory elements were determined along with several previously known motifs involved in the responses to N and carbon availability as well as plant stress. CONCLUSION: Differentially expressed genes identified provide additional insights into the coordination of the complex N responses of plants and the components of the N response mechanism. Putative N regulatory elements were identified to reveal possible new components of the regulatory network for plant N responses. A better understanding of the complex regulatory network for plant N responses will help lead to strategies to improve N use efficiency.