Oriental river prawn, Macrobrachium nipponense, is a commercially important species found in brackish and fresh waters throughout China. Chronic hypoxia is a major physiological challenge for prawns in culture, and the hepatopancreas, muscle and gill tissues play important roles in adaptive processes. However, the effects of dissolved oxygen availability on gene expression and physiological functions of those tissues of prawns are unknown. Adaptation to hypoxia is a complex process, to help us understand stress-sensing mechanism and ultimately permit selection for hypoxia- tolerant prawns, we performed transcriptomic analysis of juvenile M. nipponense hepatopancreas, gill and muscle tissues by RNA-Seq. Approximately 46,472,741; 52,773,612 and 58,195,908 raw sequence reads were generated from hepatopancreas, muscle and gill tissues, respectively. A total of 62,722 unigenes were generated, of the assembled unigenes, we identified 8,892 genes that were significantly up-regulated, while 5,760 genes were significantly down-regulated in response to chronic hypoxia. Genes from well known functional categories and signaling pathways associated with stress responses and adaptation to extreme environments were significantly enriched, including genes in the functional categories “response to stimulus”, “transferase activity” and “oxidoreductase activity”, and the signaling pathways “oxidative phosphorylation”, “glycolysis/gluconeogenesis” and “MAPK signaling”. The expression patterns of 18 DEGs involved in hypoxic regulation of M. nipponense were validated by quantitative real-time reverse-transcriptase polymerase chain reactions (qRT-PCR; average correlation coefficient = 0.94). In addition, the hepatopancreas and gills exhibited histological differences between hypoxia and normoxia groups. These structural alterations could affect the vital physiological functions of prawns in response to chronic hypoxia, which could adversely affect growth and survival of M. nipponense. Gene expression changes in tissues from the oriental river prawn provide a preliminary basis to better understand the molecular responses of M. nipponense to chronic hypoxia. The differentially expressed genes (DEGs) identified in M. nipponense under hypoxia stress may be important for future genetic improvement of cultivated prawns or other crustaceans through transgenic approaches aimed at increasing hypoxia tolerance.