Osmotic stress severely limits the growth and development of tomato (Solanum lycopersicum L.) by reducing cellular water potential, disrupting redox homeostasis, and impairing physiological functions. Micro–nano bubble (MNB) treatment can increase dissolved oxygen in the root-zone solution and improve the root-zone environment, which may benefit root metabolic activity and stress adaptation. However, the underlying molecular mechanisms are still not elucidated. To explore the underlying molecular mechanisms of how MNB-mediated root oxygenation alleviates osmotic stress in tomato, we have integrated the physiological and biochemical alterations, variable-pressure scanning electron microscopy (VP-SEM), and transcriptomic analysis (RNA-seq) under osmotic stress. The results revealed that MNBs significantly reduced PEG-induced wilting and decreased reactive oxygen species (ROS) accumulation and relative electrical conductivity (REC). Indeed, MNBs also markedly upregulated the expression of root aquaporins PIP2.7 and PIP2.4, suppressed the expression of NCED1 in leaves, and increased levels of endogenous growth-promoting hormones, including IAA and GA₃, under osmotic stress. VP-SEM observations showed that MNB-treated plants exhibited a relatively more open stomatal appearance compared with PEG-treated plants. Together, these findings suggest that MNBs mitigate PEG-induced osmotic stress in tomato, potentially by improving the root-zone aeration environment and coordinating water transport-related gene expression, antioxidant defense, and hormonal balance. These results provide a promising physical approach and theoretical basis for improving tomato stress tolerance under osmotic stress.