Objective This study preliminarily investigated the potential mechanisms of the Huoxue Tongluo prescription (HXTLP) in treating spinal cord injury (SCI) through a combination of network pharmacology, molecular docking technology, and in vivo experimental verification. Methods The traditional Chinese medicine systems pharmacology database and analysis platform (TCMSP) were utilized to select the active ingredients, targets of action were obtained from Swiss target prediction database, and an“active ingredients-targets”network was constructed. SCI-related targets were obtained by accessing online mendelian inheritance in man (OMIM) and human gene database (GeneCards), and a protein interaction network of the common targets of HXTLP and SCI was established based on the search tool for the retrieval of interacting genes/protein (STRING) database. The Metascape database was used in KEGG pathway enrichment and GO analyses of the common targets. Molecular docking of active ingredients and key targets was performed through Autodock 1.5.7 software, and the Results were visualized with Pymol 2.4.0 software. Finally, the effect of HXTLP on SCI was verified by animal experiments. Results A total of 184 intersection targets were obtained, and the key targets were serine/threonine kinase (AKT1), signal transducer and activator of transcription 3(STAT3), heat shock protein 90 kDa alpha,class A member 1 (HSP90AA1), phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1), harvey ras (HRAS), estrogen receptor 1 (ESR1), mitogen-activated protein kinase 1 (MAPK1), and epidermal growth factor receptor (EGFR). Molecular docking result showed strong binding abilities between the core active components and key targets. In the animal experiments, the behavioral scores of mice in the HXTLP group increased (P<0.05), the motor function of hind limbs was improved, and the histological morphology of the injured area was more complete compared with those of the model group. Western Blot result revealed that HXTLP effectively inhibited the key target protein (HSP90AA1) and the expression of phospho-STAT3 (P-STAT3) and promoted the expression of phospho-phosphatidylinositol-3-kinase (P-PI3K) and phospho-AKT1 (PAKT1). Conclusions This study verified that HXTLP has multi-component, multi-target, and multi-pathway synergistic effects in the treatment of SCI and has provided experimental and theoretical bases for further clinical medication research for SCI.