Objective: This study aimed to conduct proteomic analysis of the sphincter in a neurogenic bladder caused by T10 spinal cord injury. The differentially expressed proteins (DEPs) of the sphincters (internal urethral sphincter) in the neurogenic bladders (NBs) of rats after complete transection of the T10 spinal cord segment were screened using tandem mass tag (TMT)-based quantitative labeling, and their biological information was analyzed. Methods: Twelve adult Sprague Dawley rats out of 40 were randomly assigned to the blank group (n = 12), while the remaining 28 were placed in the T10 spinal cord injury model via modified Hassan Shaker spinal cord transection; 12 of these rats were then randomly selected as the model group. The rats in both groups underwent urodynamics detection and hematoxylin and eosin (H&E) staining. The proteins expressed in the bladder sphincter were detected using TMT-based quantitative proteomics. DEPs were defined as proteins with fold change >1.5 or <1/1.5, p < 0.05, and unique peptide ≥2. The DEPs were subjected to Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis using KOBAS 3.0., and gene ontology functional annotation analysis was performed using the Cytoscape 3.7.1. BiNGO plug-in. The protein-protein interaction network was then constructed using the interactive gene-retrieval tool STRING and Cytoscape software. Results: The leak-point pressure and maximum cystometric volume in the model group were significantly higher than those in the blank group (p < 0.01), and H&E staining showed continuous interruption of the bladder sphincter fibers in the model group. A total of 250 DEPs were screened in the bladder sphincter, 83 of which were upregulated and 167 of which were down-regulated. KEGG analysis of the DEPs was used to screen 15 pathways, including metabolic pathways, extracellular matrix (ECM)-receptor interaction, adhesion spots, the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway, the cytochalasin signaling pathway, and the advanced glycation end-products (AGE)/receptor for AGEs (RAGE) signaling pathway in diabetic complications and vascular smooth muscle contraction. Conclusions: It is of great significance to explore the pathological mechanism of non-inhibitory contraction of the bladder sphincter caused by spinal cord injury above the T10 segment from the perspective of ECM-receptor interaction, focal adhesion-activated PI3K/Akt signaling pathway, and cell relaxation signaling pathways. Synaptic vesicle glycoprotein (Sv2A) involved in the release of neurotransmitters from synaptic vesicles, arrestin β2 inhibitory proteins involved in α-adrenergic receptors and G-protein-coupled receptor internalization, and calmodulin and calmodulin binding protein involved in calcium-sensitive signaling pathways may be potential targets for developing new ways to treat bladder sphincter overactivity caused by T10 spinal cord injury.