Visualizing peroxynitrite dynamics in an epilepsy model using an endoplasmic reticulum-targeted fluorescent probe

Epilepsy is a chronic neurodegenerative disorder that poses a significant threat to global health. Recent studies have shown a strong correlation between the onset of epilepsy and the highly reactive nitrogen species (RNS) peroxynitrite (ONOO−), with endoplasmic reticulum (ER) stress playing a critical role in its pathological process. However, the spatiotemporal dynamics of ONOO− levels under ER stress during epilepsy remain poorly understood. To address this, we developed a novel ER-targeting fluorescent probe, ER-EO, for real-time monitoring of ONOO− changes in epileptic conditions. The ER-EO probe exhibits high specificity, precisely targeting the ER and detecting ONOO− at concentrations as low as 4.5 nM, while enabling visualization of both exogenous and endogenous ONOO−. Experimental results demonstrated that ER-EO significantly detected elevated ONOO− levels in cells under tunicamycin-induced ER stress and simulated epileptic conditions. Furthermore, in kainic acid (KA)-induced epilepsy models, including cells, zebrafish, and mice, ER-EO successfully enabled real-time monitoring of ONOO− level fluctuations during both the onset and remission phases of epilepsy. This study not only provides a powerful tool for elucidating the pathological mechanisms of ONOO− in epilepsy but also reveals the dynamic interplay between ER stress and epilepsy progression, offering new insights into the pathological processes of epilepsy and laying a foundation for the development of potential therapeutic strategies.