Abstract
PURPOSE: β-Amyrin acetate (BAA), a natural pentacyclic triterpenoid, exhibits greater lipophilicity than its antiepileptic precursor β-amyrin, implying enhanced blood-brain barrier (BBB) permeability. This study aimed to evaluate the antiepileptic efficacy of BAA and investigate its neuroprotective mechanisms, with a focus on calcium signaling. PATIENTS AND METHODS: An epilepsy model was established in zebrafish using pentylenetetrazole (PTZ) to assess the effects of BAA on seizure-like behaviors, reactive oxygen species (ROS) levels, apoptosis, and inflammatory markers. Potential targets were predicted via network pharmacology and molecular docking. Furthermore, a glutamate (Glu)-induced HT-22 neuronal injury model was used to validate BAA's effects on intracellular calcium homeostasis and downstream signaling pathways. RESULTS: BAA significantly attenuated PTZ-induced seizure-like behaviors in zebrafish, specifically reducing the frequency of clonic and tonic-clonic seizures, as well as total movement distance and velocity. Concurrently, BAA mitigated oxidative stress, apoptosis, and neuroinflammation in the zebrafish. Network pharmacology and molecular docking analyses suggested the calcium signaling pathway as a potential target, with BAA showing high binding affinity to proteins such as Bcl-2 and JAK2. In vitro experiments suggested that BAA effectively alleviated Glu-induced calcium overload in HT-22 cells. It downregulated the Bax/Bcl-2 ratio, suppressed overactivation of the JAK2/STAT3 pathway, and consequently reduced neuronal apoptosis and inflammation. The BAPTA-AM co-treatment experiment further supported that the protective effect of BAA depends on the regulation of intracellular calcium homeostasis. CONCLUSION: BAA exerts antiepileptic effects by inhibiting neuronal calcium overload and modulating the JAK2/STAT3 signaling pathway, thereby attenuating neuroinflammation and apoptosis. These findings provide a solid experimental foundation for developing BAA as a promising antiepileptic candidate and elucidate its multi-target mechanism of action.