Developing novel structural compounds with high efficacy, low neurotoxicity, and well-defined molecular targets remains a paramount objective in antiepileptic drug discovery. In this study, we designed and synthesized a series of 3-(1,2,3,6-tetrahydropyridine)-4-azaindole derivatives and evaluated their antiepileptic activity using subcutaneous pentetrazole (sc-PTZ) and maximum electric shock (MES) tests. In the sc-PTZ model, the most active compounds, 4w and 5i, exhibited median effective dose (ED50) values of 22.01 mg/kg and 25.26 mg/kg, respectively. Notably, these compounds exhibited superior safety profiles compared to standard antiepileptic drugs (AEDs) such as valproate and ethosuximide, both compounds demonstrated lower neurotoxicity and higher protective indexes (PI = TD50/ED50) with PI values exceeding 27.26 and 23.75, respectively. Detailed structure-activity relationship (SAR) studies indicated that the N-atom at the 4-position and the H-atom of the NH unit in the 4-azaindole skeleton, and the double bond in 1,2,3,6-tetrahydropyridine are critical for their antiepileptic activities. Mechanistic results revealed that both compounds 4w and 5i primarily target voltage-gated sodium channels, rather than GABAA or NMDA receptors. Further studies indicated that compounds 4w and 5i effectively block sodium ion channels and significantly inhibit Nav1.2 at the cellular level, which was further supported by molecular docking of compound 4w and 5i with Nav1.2 (PDB ID: 6J8E) receptor in silico.
