Roots play important roles in anchoring plants in the soil and absorbing water and nutrients from the soil. Soil salinity is one of the major abiotic stresses that adversely affect root growth and development. The exogenous application of some bioactive substances is a simple and effective approach to improve plant stress tolerance in agricultural production. Nitric oxide (NO) is a bioactive, gaseous, multifunctional molecule which plays a central role and mediates a variety of physiological processes and responses to biotic and abiotic stresses, including salt. However, information on the effects of exogenous SNP on alfalfa root systems under salt stress conditions and its physiological mechanisms is lacking. In this study, Medicago sativa L. cv. Gannong No.4 was used as the experimental material and NO-donor sodium nitroprusside (SNP), NO-scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO), tungstate, the nitrate reductase (NR) inhibitor, nitric oxide synthase (NOS) inhibitor NG-nitro-L-Arg-methyl ester (L-NAME) and sodium ferrocyanide (SNP analogue, does not release NO) were used to investigate the effects of NO on growth, root vigor, osmoregulatory molecules, membrane lipid peroxidation, reactive oxygen species (ROS) accumulation and the activity of antioxidant enzyme in alfalfa roots under NaCl stress. Results showed that when 100 μmol/L SNP was applied in the nutrient solution under NaCl stress, the growth of alfalfa roots was significantly improved for eight days, the root vigor value and the content of free proline were increased and the content of soluble protein was decreased throughout the treatment period. SNP also caused an increase in the activities of ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), guaiacol peroxidase (GPX), superoxide dismutase (SOD), and the contents of reduced ascorbic acid (AsA) and reduced glutathion (GSH) under salt stress. Whereas, the content of H₂O₂, OH^·, production rate of O₂^·- and MDA level were decreased in alfalfa roots. Meanwhile, SNP pretreatment improved the endogenous NO accumulation. Sodium ferrocyanide, the NO-donor SNP analogue, has no noticeable effect on the damage caused by NaCl stress for alfalfa roots. When applied with the NO-scavenger c-PTIO, under NaCl stress, the NR inhibitor tungstate and NOS inhibitor L-NAME both reduced the accumulation of endogenous NO in roots, further inhibited the activity of the antioxidant system, aggravated membrane lipid peroxidation and root growth, however, their harmful effects on growth, antioxidant system and membrane lipid peroxidation could be reversed by the addition of SNP. Thus, the results showed that application of SNP significantly increased root vigor, activated the antioxidant system, alleviated the oxidative root damage induced by salt stress and promoted root growth under NaCl stress. Endogenous NO may also take part in the regulation of salt-tolerance of alfalfa roots under NaCl stress, and the release of NO according to the NOS and NR pathways may play an important role in alleviating the inhibitive effect in alfalfa root growth. These results may provide a theoretical basis for the mechanisms of salt resistance, help research into NO in the chemical regulation of alfalfa salt tolerance, the breeding of salt tolerant alfalfa cultivars, and the utilization of saline land in the future.