The response to salinity may largely vary not only among species but even cultivars or lines of the same species. Knowledge of the reasons underlying these differential responses can be critical in breeding programs to obtain lines with enhanced performance under salinity. In this work, the responses to salt stress of three Medicago truncatula lines with contrasting salt resistance, TN6.18 (sensitive), Jemalong (reference line), and TN8.20 (resistant), have been assessed by analyzing a full array of nodule parameters (water relations, carbohydrates, ion concentrations, and enzyme activities). The aim of this work was to look for the most important criteria conferring resistance to the M. truncatula-Sinorhizobium symbiosis under salinity. The resistance of M. truncatula to salt stress was related to nodule osmotic adjustment due to both sequestration of sodium and accumulation of soluble carbohydrates and free amino acids following protein degradation, together with an adequate nitrogen metabolism due to maintaining relatively high glutamine synthetase activity (GS) and stimulation of NADH-dependent glutamate dehydrogenase (GDH). Glutamine synthetase activity differed clearly between the three studied lines. Thus, it may have a key role in the resistance of Medicago truncatula to salt stress.