Leaf intracellular water is the retained part of transpiration water when it flows through leaf mesophyll cells, the intracellular water is directly and closely related to photosynthesis and growth of plant. However, little is known about the dynamic use traits of intracellular water and the influence on instantaneous water-use efficiency (WUE_i) of plants at different water conditions. In this study, tomato (Solanum lycopersicum L.) plants were subjected to three different water supply strategies by regulating the soil relative water content (SWC_R) (i.e., T1: 70 %–80 %–90 %, T2: 80 %–90 %–100 %, T3: 60 %–70 %–80 %) within three treatment phases (P1, P2 and P3). The electrophysiological and photosynthetic parameters, leaf water potential, nutrient contents, growth indices and yield were determined. Leaf intracellular water use traits including transport rate (LIWTR), water-holding capacity (LIWHC) and water-use efficiency (LIWUE) were calculated according to the Nernst equation using plant electrophysiological parameters. The results showed that photosynthesis, growth and yield of tomatoes could be promoted by increasing the water supply. Plants at T3 treatment initially experienced drought-hardening and then could adapt to the surroundings and maintain high WUE_i with increasing water supply at the following phases. Besides, the plants at T3 treatment only showed a small amount (9 %) of yield loss compared to control. High value of LIWTR and low value of LIWHC indicated that less water supply could facilitate the water transport within leaf cells, which improved the WUE_i rather than the LIWUE. Sufficient water supply promoted the transpiration but did not accelerate the water transport within leaf cells and caused low value of WUE_i. 70 %–80 % SWC_R was a turning point for the changing status of leaf intracellular water in plants. In this study, the water supply strategy at T3 treatment was more conducive to balance the WUE improvement and yield loss in tomato plants than the other two. The use traits of leaf intracellular water based on plant electrophysiological parameters could provide support for the quick evaluation of plant water status.