Potassium (K) is the most required macronutrient by Eucalyptus, while sodium (Na) can partially substitute some physiological functions of K and have a positive response on plant growth in K-depleted tropical soils. However, the right percentage of K substitution by Na is not yet known for Eucalyptus seedlings, since a few experiments have only compared treatments receiving K or Na. This study evaluated five levels of Na supply (0, 0.45, 0.90, 1.35 and 1.80 mM) as substitution for K in Eucalyptus seedlings grown in nutrient solution. Plants growth, biomass, K-nutritional status, leaf gas exchange, leaf carbon isotopic composition (δ13C ‰), leaf water potential (Ψw), leaf area (LA), stomatal density (SD) and water use efficiency (WUE) were measured. The highest total biomass yield was achieved by the Na estimated rate of 0.25 mM, corresponding to a leaf K: Na ratio of 3.41, and having the lowest δ13C values. Conversely, the highest Na rate (1.8 mM) induced K deficiency symptoms, lower growth, reduced total dry matter yield, leaf gas exchange, LA, SD and a higher δ13C, which presented a trend to an inverse correlation with CO2 assimilation rate (A), WUE and shoot dry matter. Collectively, our results conclude that substitution of 25% of K by Na (0.45 mM of Na) provided significant gains in nutritional status and positive plant physiological responses by increasing WUE, stomatal diffusion, and by augmenting CO2 uptake efficiency. This nutritional management can therefore be an alternative option to optimize yields and resource use efficiencies in Eucalyptus cultivation.