BACKGROUND: Neural stem cells (NSCs) are known for their high capacity for proliferation, self-renewal, and multidirectional differentiation. However, they often fail to survive or differentiate into mature neurons capable of replacing lost neurons. This study evaluated the potential role of low-intensity pulsed ultrasound (LIPUS) in promoting NSC proliferation and differentiation both in vitro and in vivo, as well as its role in enhancing spinal cord injury (SCI) recovery. METHODS: NSCs were isolated, stimulated with LIPUS, and characterized through identification and detection assays. The safety and efficacy of LIPUS in promoting NSC proliferation and differentiation were assessed through cell viability and apoptosis assays, and neuronal marker expression analysis. In vivo, NSCs encoding fluorescent proteins were transplanted into a rat model of SCI. The SCI rats received LIPUS for 4 weeks. Later, functional recovery, morphological changes and neuronal structures were evaluated. RESULT: The isolated NSCs were successfully identified. LIPUS significantly enhanced NSC proliferation and increased the expression of key neurogenic markers and neurotrophic factors, while reducing GFAP expression and avoiding apoptosis. In vivo, the NSCs/LIPUS + group demonstrated higher survival and differentiation of transplanted NSCs, along with improved BBB scores and enhanced neural marker expression compared with the NSCs/LIPUS - group. CONCLUSION: LIPUS stimulation effectively promoted NSC proliferation and differentiation and enhanced the survival and function of transplanted NSCs at the SCI site, leading to improved SCI recovery.