To study the response mechanism of Macrobrachium nipponense to high-temperature stress, we set up a control (20±0.5) ℃ and high-temperature (30±0.5) ℃ groups. The dissolved oxygen concentration was (6.5±0.5) mg/L. The expression profiles of heat shock proteins 21 (MnHSP21), 60 (MnHSP60), 70-3 (MnHSC70-3), and 90 (MnHSP90) in M. nipponense hepatopancreas and gill tissues were measured at 0, 12, 24, 36, and 48 h under high-temperature stress. Then we measured the activities of superoxide dismutase (SOD), glutathione S-transferase (GST), glutathione peroxidase (GPX), and catalase (CAT), as well as tissue structure of M. nipponense under high-temperature stress. The results showed that four heat shock protein genes were induced in hepatopancreas and gill tissues. The expression of MnHSC70-3 gene was the most significant among the four heat shock protein genes, indicating that this gene is an important molecule for high-temperature stress response in M. nipponense. At the same time, expression trends of the four genes were more obvious in the hepatopancreas than in gill tissue, indicating that hepatopancreas is a more important organ for M. nipponense in terms of high-temperature stress response. Under high-temperature stress, the activity of all related antioxidant enzymes changed to varying degrees. The activity of SOD, GST, and CAT in the hepatopancreas increased significantly, while the activity of GPX changed only slightly. The SOD enzyme activity in gill tissue was significantly lower than in the control group (P＜0.05), and the enzyme activity levels of GST, GPX, and CAT were significantly higher than in the control group (P＜0.05). The structure of hepatopancreas and gill tissue changes under high-temperature stress. The secretory cells and internal transport vesicles in hepatopancreas increase in volume, while the layered epithelium of the gill tissue structure was slightly curved, and the arrangement of blood cells was disordered. Compared with 36 h, there was no obvious change in tissue structure after 48 h of stress. In summary, high-temperature stress activated the antioxidant system of M. nipponense and induced the expression of heat shock protein genes. The MnHSC70-3 gene may play an important role in the heat tolerance of M. nipponense. Changes in the activity of antioxidant enzymes under high-temperature stress may be essential to avoid oxidative damage. This study aimed to provide a reference for healthy farming by clarifying the response mechanism of M. nipponense to hightemperature stress.