Status epilepticus (SE), a prolonged seizure lasting over 5 minutes, is a neurological emergency associated with high morbidity and mortality. SE causes neuronal death, primarily through excitotoxicity from excessive calcium influx. Mitochondrial dysfunction plays a crucial role, impacting energy production (ATP), reactive oxygen species (ROS) homeostasis, and calcium buffering. Mitochondrial dynamics, the balance of fission (fragmentation) and fusion (elongation), are critical for neuronal survival. Impaired mitochondrial dynamics, particularly excessive fusion (hyperfusion), leading to ROS production and ATP deficiency, contribute to SE-induced neuronal death. Epigallocatechin-3-gallate (EGCG), a major component of green tea, possesses antioxidant properties and has shown neuroprotective effects in various models. While EGCG protects hippocampal neurons from SE, the underlying mechanisms remain unclear. This study aimed to investigate the role of EGCG in regulating mitochondrial dynamics and related signaling pathways in SE-induced CA1 neuronal degeneration. The study hypothesized that EGCG's neuroprotection against SE involves the modulation of mitochondrial dynamics through specific signaling pathways, including GPx1, ERK1/2, DRP1, and NF-κB.

The literature extensively documents the detrimental effects of SE on neuronal survival and function, highlighting the crucial role of excitotoxicity and mitochondrial dysfunction. Studies have shown that aberrant mitochondrial dynamics, particularly hyperfusion, contribute to neuronal death in SE models. Previous research has demonstrated EGCG's antioxidant and neuroprotective properties in various neurological conditions. However, the specific mechanisms by which EGCG protects against SE-induced neuronal damage and its impact on mitochondrial dynamics remain incompletely understood. The existing literature suggests potential links between EGCG, oxidative stress, mitochondrial function, and signaling pathways involved in neuronal survival and inflammation.

Seventy male Sprague-Dawley rats were divided into three groups: control, EGCG (50 µM), and EGCG (50 µM) + U0126 (25 µM, ERK1/2 inhibitor). SE was induced using lithium chloride and pilocarpine. Three days post-SE, animals were sacrificed, and hippocampal tissues were collected for various analyses. Western blotting was used to assess the protein levels of GPx1, DRP1, p-DRP1 (S616), ERK1/2, p-ERK1/2, JNK, and p-JNK. Immunohistochemistry and mitochondrial morphometry were performed to analyze mitochondrial morphology (elongation index and aggregation) and GPx1, DRP1, and p-ERK1/2 expression in CA1 neurons. Fluoro-Jade B (FJB) staining quantified neuronal degeneration. Statistical analysis involved Mann-Whitney or Kruskal-Wallis tests with Dunn-Bonferroni post hoc comparisons.

EGCG significantly attenuated SE-induced CA1 neuronal death (p=0.009). EGCG prevented SE-induced GPx1 downregulation in CA1 neurons (p<0.001). EGCG significantly reduced SE-induced mitochondrial elongation and aggregation in CA1 neurons (p<0.001). EGCG prevented SE-induced decreases in DRP1 expression and its S616 phosphorylation in CA1 neurons (p<0.001 and p=0.01 respectively). EGCG increased ERK1/2 phosphorylation but did not affect JNK phosphorylation following SE (p=0.009 for ERK1/2 phosphorylation). EGCG decreased NF-κB S536 phosphorylation in CA1 neurons following SE (p<0.001). Co-treatment with U0126 abolished EGCG's neuroprotective effect (p=0.015), increased mitochondrial elongation and aggregation, and blocked EGCG-induced DRP1 S616 phosphorylation (p=0.013), without affecting GPx1 or NF-κB levels.

The findings demonstrate that EGCG's neuroprotective effects against SE are mediated, at least in part, by its actions on GPx1, ERK1/2-DRP1, and NF-κB pathways. EGCG's antioxidant properties, leading to GPx1 upregulation, likely contribute to the reduction of oxidative stress and the preservation of mitochondrial function. The enhancement of ERK1/2-DRP1-mediated mitochondrial fission by EGCG suggests a crucial role for balanced mitochondrial dynamics in neuroprotection. The inhibition of NF-κB S536 phosphorylation by EGCG points to a role for EGCG in reducing inflammation in response to SE. The pivotal role of ERK1/2 activation in EGCG's neuroprotective effects is strongly supported by the complete reversal of EGCG’s protective effects by U0126, indicating a direct link between ERK1/2 activation and EGCG’s neuroprotective mechanism.

This study demonstrates that EGCG protects CA1 neurons from SE-induced death through multiple mechanisms involving GPx1 upregulation, ERK1/2-dependent restoration of mitochondrial fission, and inhibition of NF-κB activation. The results highlight the therapeutic potential of EGCG in mitigating SE-induced neuronal damage. Future research could focus on further elucidating the molecular interactions within these pathways and exploring potential clinical applications of EGCG in epilepsy treatment. Investigation into EGCG's effects on hypoxia-related events during SE, given its poor blood-brain barrier permeability, is also warranted.

The study utilized an intracerebroventricular administration of EGCG due to its poor blood-brain barrier permeability. This limits the generalizability of the findings to in vivo scenarios where EGCG is administered peripherally. The study did not directly investigate the effects of hypoxia during SE, despite the potential for hypoxia to contribute to hippocampal damage. Further research is needed to fully clarify EGCG's effects in more physiologically relevant settings with different administration routes and assessing the potential impact of hypoxia.