Abstracts

Rationale: The adipocyte-derived hormone leptin plays a crucial role in regulating body weight and metabolism by activating leptin receptors. Emerging evidence suggests that leptin receptors are also expressed in microglia, the resident immune cells of the central nervous system (CNS), where they may influence neural pathophysiological processes. Recent studies indicate that leptin is implicated in the regulation of NLRP3 inflammasome activation. However, the underlying mechanisms are largely unknown. Here in this study, we sought to determine whether leptin regulates microglial NLRP3 activation and consequently affect seizure activity.


Methods: We established a transwell neuron-microglia co-culture system to investigate the impact of leptin on microglial NLRP3 activation in epileptic conditions. Microglia, co-cultured with magnesium (Mg2+)-free medium-treated neurons, underwent immunohistochemical and biochemical assays to assess NLRP3 activation and leptin receptor expression. Whole-cell current clamp recordings were conducted on microglia-co-cultured Mg²⁺-free medium-treated neurons to evaluate neuronal epileptiform activity. In the in vivo experiment, we utilized a pilocarpine-induced temporal lobe epilepsy (TLE) mice model to study the effect of leptin on microglial NLRP3 activation. Seizure frequency and severity were assessed using a video EEG monitoring system. Single-cell RNA sequencing (scRNA-seq) and transcriptome analysis were employed to elucidate the molecular mechanisms underlying leptin-mediated NLRP3 activation in microglia of epileptic mice.


Results: Our findings demonstrate that microglial NLRP3 inflammasome activation is enhanced, and leptin receptor expression is upregulated when co-cultured with Mg²⁺-free medium-treated neurons. Activation of microglial leptin receptors induces NLRP3 inflammasome activation, while blocking these receptors suppresses NLRP3 inflammasome activation. Furthermore, whole-cell current clamp recordings reveal a significant decrease in the frequency of spontaneous action potentials (sAPs) in leptin receptor-deficient microglia-co-cultured epileptiform neurons compared to control microglia-co-cultured neurons. Consistent with our in vitro results, NLRP3 inflammasome activation and leptin receptor expression are increased in the TLE mice model. Depletion of leptin receptors suppresses NLRP3 inflammasome activation and seizure activity. scRNA-seq and transcriptome analysis identify galectin-1 (Lgals1) from the galectin family as a potential key player in microglial leptin-mediated NLRP3 activation in epilepsy. Microglial activation, followed by seizure activity, significantly enhances Lgals1 expression in a leptin-dependent manner, which is crucial for NLRP3 inflammasome activation.


Conclusions: In conclusion, our results suggest that leptin drives microglial NLRP3 inflammasome activation, leading to increased seizure activity. This study sheds light on the intricate interplay between leptin, microglia, and the NLRP3 inflammasome in epilepsy, highlighting potential therapeutic targets for epilepsy treatment.


Funding:
This work was supported by grants from the National Natural Science Foundation of China (81872846 and 81673413 to Xinjian Zhu).